Http://2014.igem.org/Team:Heidelberg/pages/Notebook/Materials

From 2014.igem.org

(Difference between revisions)
(→Kits)
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| 2x PCR Master mix Solution (iTaq) || HISS DIAGNOSTICS GmbH || 25028
| 2x PCR Master mix Solution (iTaq) || HISS DIAGNOSTICS GmbH || 25028
 +
|}
 +
<br/>
 +
 +
=Antibodies=
 +
{|class="wikitable sortable"
|-
|-
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| Pronase from ''Streptomyces griseus'' || Sigma-Aldrich Chemie GmbH || P6911-100MG
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! Antibody !! Supplier !! Catalog Number
 +
|-
 +
| Penta·His Antibody, BSA-free || QIAGEN || 34660
 +
|-  
 +
| Anti-GFP || Roche || 11814460001
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|-
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| mRFP
 +
|-
 +
| Goat anti Mouse IgG || HRPO Dianova GmbH || 115-035-003
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|-
 +
| secondary rat
|}
|}
<br/>
<br/>
 +
=Restriction Enzymes=
=Restriction Enzymes=
{|class="wikitable sortable"
{|class="wikitable sortable"
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<br/>
<br/>
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=Miscellaneous Primers=
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=Primers and Oligos=
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{| class="wikitable sortable"
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|-
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! Identifier !! Order date !! Note !! Sequence
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|-
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| VF2 || - || for screening in standard BB backbone- binds on the Backbone before Insert  || TGCCACCTGACGTCTAAGAA
+
-
|-
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| VR || - || for screening in standard BB backbone- binds on Backbone behind Insert || ATTACCGCCTTTGAGTGAGC
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|-
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-
|}
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<br/>
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=Primers and Oligos=
 
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==Delftibactin==
 
{| class="wikitable sortable" style="width:50%;"
{| class="wikitable sortable" style="width:50%;"
|-
|-
! Identifier !! Order date !!Note !! Sequence
! Identifier !! Order date !!Note !! Sequence
-
|-
 
-
| DN01:delH_f1_PacI_fw || - || fw Primer for DelH-fragment1 with RBS and PacI-restriction site || TTTT TTAATTAA    TCACACAGGAAAGTACTAG<br/>ATGGACCGTGGCCGCCTGC    GCCAAATCG
 
-
|-
 
-
| DN02:delH_f1_SalI_rev || - || - || TTTT GTCGACCAACACCTGTGCCTGC
 
-
|-
 
-
| DN03:delH_f2_SalI_fw || - || - || TTTT GTCGACTGGATGGAGCCTGGTGAAAG
 
-
|-
 
-
| DN04:delH_f2_KpnI_rev || - || - || TTTT GGTACC  TCAGTCCAGCGCGTACTCCAG
 
-
|-
 
-
| DN05:AraCbb_KpnI_fw || - || amplifying the Backbone for DelH (pSB6A1-AraC-lacZ) || TTTT  GGTACC  AAAGAGGAGAAATACTAGATGACCATG
 
-
|-
 
-
| DN06:AraCbb_PacI_rev2 ||  15-05-2013  || amplifying the Backbone for DelH (pSB6A1-AraC-lacZ) || TTTT  TTAATTAA  GCTAGCCCAAAAAAACGGTATG
 
-
|-
 
-
| DN07:Screen_delH_rev|| 15-05-2013 || for screening if DelH is present - binds on the very beginning of DelH || CTTTCCTCGAACACCGTGCGCAG
 
-
|-
 
-
| DN08:DelH_EcoRI_rev || - || rev_Primer for DelH Fragment f1a || CTCGTCGCCATGGACCAGGCAG
 
-
|-
 
-
| DN09:DelH_f1_fw_long || 2013-06-11 || for amplifying DelH-1a from the genome: doesn't work|| ATGGACCGTGGCCGCCTGCGCCAAATCG
 
-
|-
 
-
| DN10:DelH_f1_fw_short || 2013-06-11 || for amplifying DelH-1a from the genome: doesn't work || ATGGACCGTGGCCGCCTGC
 
-
|-
 
-
| DN11:DelH_f1_fw_short2|| 2013-06-11 || for amplifying DelH-1a from the genome: works!!!! || GCCGCCTGCGCCAAATCG
 
-
|-
 
-
| DN12:DelH_f1_PacI_fw_short|| 2013-06-11 || for amplifying DelH-1a from the genome: doesn't work || TTTTTTAATTAATCACACAGGAAAGTAC<br/>TAGATGGACCGTGGCCGCCTGC
 
-
|-
 
-
| DN13:Screen_DelH_fw|| 15-05-2013 || PCR Screening for presence of DelH insert || GTAAACCCACTGGTGATACCATTC
 
-
|-
 
-
| FS_01: pSB4K5_DelA_rv||20-13-06-28||Amplification of pSB4K5 from the iGEM Distribution Gibson Primer with overhang to DelA introducing the RBS BBa_B0035|| TCGCGGCGATCCGGTACTGCGCCTCTGTT<br/>GAACATCTGATATTCTCCTCTTTAATCG<br/> ACAGATTGTGTGAAATTGTTATCCGCTCAC
 
-
|-
 
-
|FS_02: DelAG_1_fw||2013-06-28 || Amplification of DelAG from ''Delftia acidovorans'' genome Gibson Primer|| TTCAACAGAGGCGCAGTACCGGATC
 
-
|-
 
-
|FS_03: DelAG_1_rv||2013-06-28 ||Amplification of DelAG from ''Delftia acidovorans'' genome Gibson Primer|| GTCGGAGACGATGTGGTGCATCAC
 
-
|-
 
-
|FS_04: DelAG_2_fw||2013-06-28 ||Amplification of DelAG from ''Delftia acidovorans'' genome Gibson Primer|| CTGCAGGCCAATGAGCACATCCTG
 
-
|-
 
-
|FS_05: DelAG_2_rv||2013-06-28 ||Amplification of DelAG from ''Delftia acidovorans'' genome Gibson Primer|| CACAGGTGGTAGATGGCGTC
 
-
|-
 
-
|FS_06: DelAG_3_fwG||2013-06-28 ||Amplification of DelAG from ''Delftia acidovorans'' genome Gibson Primer|| ATTGCGAGGACTTGCTCGATG
 
-
|-
 
-
|FS_07: DelAG_3_rv||2013-06-28 ||Amplification of DelAG from ''Delftia acidovorans'' genome Gibson Primer|| TTTGCTGCAGCGCCAGCACATCGAG
 
-
|-
 
-
|FS_08: DelAG_4_fw||2013-06-28 ||Amplification of DelAG from ''Delftia acidovorans'' genome Gibson Primer|| GTACGGCCTATCACATCAGCG
 
-
|-
 
-
|FS_09: DelAG_4_rv||2013-06-28 ||Amplification of DelAG from ''Delftia acidovorans'' genome Gibson Primer|| GAAGCTCAGCAGGTTGGGCGAGACG
 
-
|-
 
-
|FS_10: DelAG_5_fw||2013-06-28 ||Amplification of DelAG from ''Delftia acidovorans'' genome Gibson Primer|| GAATTTTGTTCCACCACCTGCTG
 
-
|-
 
-
|FS_11: DelAG_5_rv||2013-06-28 ||Amplification of DelAG from ''Delftia acidovorans'' genome Gibson Primer with overhang to DelOP|| CTTGAGCAGGCGCAGTACCTCGGAGGG<br/> CGGTCGGCTGGCGTTTTCCATGATT<br/>CAGGTTTCCTGTGTGAAGCTCATCTCAGATA<br/> TCTCCCAGAGTTTCGAGAAAG
 
-
|-
 
-
|FS_11: DelAG_5_short_rv||2013-05-07 ||Amplification of DelAG from ''Delftia acidovorans'' genome Gibson Primer|| TCAGATATCTCCCAGAGTTTCGAGAAAG
 
-
|-
 
-
|FS_12: DelOP_fw||2013-06-28 ||Amplification of DelOP from ''Delftia acidovorans'' genome Gibson Primer|| GAATCATGGAAAACGCCAGCCGAC
 
-
|-
 
-
|FS_13: DelOP_rv||2013-06-28 ||Amplification of DelOP from ''Delftia acidovorans'' genome Gibson Primer with overhang to DelL|| CAATGTTGGAGGGGCCGAAGCCGATGCCGATC<br/>AGCGGGTGGGTTTGCATGGAAGGTC CTTTCATTGGGTCGATGCGTCCAGTGT<br/> CACACCGTGGTGTCTGCAGGCG
 
-
|-
 
-
|FS_13: DelOP_short_rv||2013-05-07 ||Amplification of DelOP from ''Delftia acidovorans'' genome Gibson Primer with overhang to DelL||TCACACCGTGGTGTCTGCAGGCG
 
-
|-
 
-
|FS_14: DelL_fw||2013-06-28 ||Amplification of DelL from ''Delftia acidovorans'' genome Gibson Primer|| CAAACCCACCCGCTGATCGGCATC
 
-
|-
 
-
|FS_15: DelL_mRFP_pSB4K5_rv||2013-06-28 ||Amplification of DelL Gibson Primer with overhang to BBa_J04450|| GAAACGCATGAACTCTTTGATAACGTCT TCGGAGGAAGCCAT<br/>CTAGTATTTCTCCT CTTTCTCTAGTATCAGTCCTGCAGCG CCAGCTGTTCTGTG
 
-
|-
 
-
|FS_15: DelL_mRFP_pSB4K5__short_rv||2013-05-07 ||Amplification of DelL Gibson Primer with overhang to BBa_J04450||TCAGTCCTGCAGCGCCAGCTGTTCTGTG
 
-
|-
 
-
|FS_16: mRFP_pSB4K5_fw (Del)||2013-06-28 ||Amplification of pSB4K5 from iGEM Distribution Gibson Primer|| GCTTCCTCCGAAGACGTTATC
 
-
|-
 
-
|FS_20: DelF_fw||2013-07-13 ||Amplification of DelF from ''Delftia acidovorans'' genome Gibson Primer|| GACTTGCTCGATGCGGTGCAG
 
-
|-
 
-
|FS_21: DelF_fw||2013-07-13 ||Amplification of DelF from ''Delftia acidovorans'' genome Gibson Primer|| GACGCCATCTACCACCTGTG
 
-
|-
 
-
|FS_22: DelOP_short_fw||2013-08-02 ||Amplification of DelOP from ''Delftia acidovorans'' genome inlcuding the recently predicted endogenous Promotor for DelOP Gibson Primer|| GATGACGCAGGGCGGCGGAATTTGTTCATC
 
-
|-
 
-
|FS_23: DelG_long_rv||2013-07-13 ||Amplification of DelG from ''Delftia acidovorans'' genome Gibson primer with overhang to DelOP element including the recently predicted endogenous Promotor|| GATGAACAAATTCCGCCGCCCTGCGTCA<br/>TCTCAGATATCTCCCAGAGTTTCGAGAAAG
 
-
|-
 
-
|FS_24: DelAE_rv||2013-07-13 ||Amplification of DelAE from ''Delftia acidovorans'' genome Gibson Primer|| CAGAAGAATTCCCAGAAGGAGATGTCGAAG
 
-
|-
 
-
|FS_25: DelEF_fw||2013-07-13 ||Amplification of DelEF from ''Delftia acidovorans'' genome Gibson Primer|| ACACGGTGCTGCAGAAAACGCCCTTC
 
-
|-
 
-
|FS_26: DelFG_rv||2013-07-13 ||Amplification of DelFG from ''Delftia acidovorans'' genome Gibson Primer|| GAATTCATCCACGATGATCTGCATG
 
-
|-
 
-
|FS_27: DelOP_rv||2013-08-02 ||Amplification of DelOP from ''Delftia acidovorans'' genome Gibson Primer|| CTTTGGGCCGTGCCGGTTTTTGAGATAC
 
-
|-
 
-
|FS_28: DelOP_rv||2013-08-02 ||Amplification of DelOP from ''Delftia acidovorans'' genome Gibson Primer|| GTTTTTGAGATACGCGCGTTGTCAC
 
-
|-
 
-
|FS_29: DelOP_rv||2013-08-02 ||Amplification of DelOP from ''Delftia acidovorans'' genome Gibson Primer|| TTCCCCCTCTCTTTCTCGCTTC
 
-
|-
 
-
|FS_30: DelOP_rv||2013-08-02 ||Amplification of DelOP from ''Delftia acidovorans'' genome Gibson Primer|| CCGCTTCCCCCTCTCTTTCTCGCTTC
 
-
|-
 
-
|FS_31: DelOP_fw||2013-08-02 ||Amplification of DelOP from ''Delftia acidovorans'' genome Gibson Primer|| GTTGGCGAGTTCAAGAAATG
 
-
|-
 
-
|FS_32: DelOP_fw||2013-08-02 ||Amplification of DelOP from ''Delftia acidovorans'' genome Gibson Primer|| TCCTTCAGGTGTGCGGCAGACAAG
 
-
|-
 
-
|FS_33: DelOP_fw||2013-08-02 ||Amplification of DelOP from ''Delftia acidovorans'' genome Gibson Primer|| TTCTTCGTGATGACGCAGGGCGGCGGAATTTGTTC
 
-
|-
 
-
|FS_35: DelG_fw||2013-08-05 ||Amplification of DelG from ''Delftia acidovorans'' genome Gibson Primer|| CATGCAGATCATCGTGGATGAATTC
 
-
|-
 
-
|FS_45: pSB4K5_fw||2013-08-02 ||Amplification of pSB4K5 from iGEM Distribution Gibson Primer without mRFP|| CCAGGCATCAAATAAAACGAAAG
 
-
|-
 
-
|FS_46: DelL_rv ||2013-08-02 ||Amplification of DelL from ''Delftia acidovorans'' genome Gibson Primer creating overlap to pSB4K5 without mRFP|| TCAGTCCTGCAGCGCCAGCTGTTCTG TGCTTTCGTTTTATTTGATGCCT
 
-
|-
 
-
|FS_47_screening_BB_AF_fw||2013-08-02 ||Primer for screening/sequencing of pFSN construct|| GTTGGCCGATTCATTAATGC
 
-
|-
 
-
|FS_48_screening_BB_AF_rv||2013-08-02 ||Primer for screening/sequencing of pFSN construct|| TAACGGTATCGGTATCGCTTTG
 
-
|-
 
-
|FS_49_screening_AFI_AFII_fw||2013-08-02 ||Primer for screening/sequencing of pFSN construct|| GTTTCTCTGGAAGATGGATAC
 
-
|-
 
-
|FS_50_screening_AFI_AFII_rv||2013-08-02 ||Primer for screening/sequencing of pFSN construct|| GTTGACGAAAAAGCCGACCAC
 
-
|-
 
-
|FS_51_screening_AF_FG(21-26)_fw||2013-08-02 ||Primer for screening/sequencing of pFSN construct|| TGGATATCGACTGGACTGCCTG
 
-
|-
 
-
|FS_52_screening_AF_FG(21-26)_rv||2013-08-02 ||Primer for screening/sequencing of pFSN construct|| TGCACCACATCGACGAAACGG
 
-
|-
 
-
|FS_53_screening_FG(21-26)_G_fw||2013-08-02 ||Primer for screening/sequencing of pFSN construct|| GTACGGCCTATCACATCAGCG
 
-
|-
 
-
|FS_54_screening_FG(21-26)_G_rv||2013-08-02 ||Primer for screening/sequencing of pFSN construct|| GAACCTGGGTGTTCACGAAAAAGCC
 
-
|-
 
-
|FS_55_screening_G_OP_fw||2013-08-02 ||Primer for screening/sequencing of pFSN construct|| GTATCTCTACATGCATCGCTAC
 
-
|-
 
-
|FS_56_screening_G_OP_fw||2013-08-02 ||Primer for screening/sequencing of pFSN construct|| AGGACATTTTCCGCACCCCG
 
-
|-
 
-
|FS_57_screening_G_OP_rv||2013-08-02 ||Primer for screening/sequencing of pFSN construct|| GCTGGCGTTTTCCATAAG
 
-
|-
 
-
|FS_58_screening_OP_L_fw||2013-08-02 ||Primer for screening/sequencing of pFSN construct|| GAACAACTTCCAGCACAGCCTGTTC
 
-
|-
 
-
|FS_59_screening_OP_L_rv||2013-08-02 ||Primer for screening/sequencing of pFSN construct|| CGTTGAAGATTTCGTTGACG
 
-
|-
 
-
|FS_60_screening_L_BB_fw||2013-08-02 ||Primer for screening/sequencing of pFSN construct|| CATCTTCAAGGTGTTCTATGAAC
 
-
|-
 
-
|FS_61_screening_L_BB(with_mRFP)_rv||2013-08-02 ||Primer for screening/sequencing of pFSN construct|| CAGTTTAACTTTGTAGATGAAC
 
-
|-
 
-
|FS_66: DelH_rv||2013-08-26 || Amplification of DelH from ''Delftia acidovorans'' genome Gibson Primer|| TGGGCATTCACCGCATCGATC
 
-
|-
 
-
|FS_67: DelH_fw||2013-08-26 || Amplification of DelH from ''Delftia acidovorans'' genome Gibson Primer|| CTTCACGTTGATTGCGCATG
 
-
|-
 
-
|FS_68: DelH_rv||2013-08-26 || Amplification of DelH from ''Delftia acidovorans'' genome Gibson Primer|| CAGAAGAACTCCCAGACCGAC
 
-
|-
 
-
|FS_69: DelH_fw||2013-08-26 || Amplification of DelH from ''Delftia acidovorans'' genome Gibson Primer|| GACACCGTTCAGCTTCGATG
 
-
|-
 
-
|FS_70: DelH_rv||2013-08-26 || Amplification of DelH from ''Delftia acidovorans'' genome Gibson Primer|| GAAGCTGCTCCGCTGATAGAT
 
-
|-
 
-
|FS_71: DelH_fw||2013-08-26 || Amplification of DelH from ''Delftia acidovorans'' genome Gibson Primer|| ATGTGCTGTCGCTCAAGATG
 
-
|-
 
-
|FS_72_SR_02_fw||2013-08-30 || Screening of pFHFSN|| ATGTGCTGTCGCTCAAGATG
 
-
|-
 
-
|FS_73_SR_03_fw||2013-08-30 || Screening of pFHFSN|| GTGCTGTTTGGCCGTATG
 
-
|-
 
-
|FS_74_SR_04_fw||2013-08-30 || Screening of pFHFSN|| ATCAGGTGCTGAGCTACGAC
 
-
|-
 
-
|FS_75_SR_05_fw||2013-08-30 || Screening of pFHFSN|| CTGTTCATCAACACCTTGCC
 
-
|-
 
-
|FS_76_SR_06_rv||2013-08-30 || Screening of pFHFSN|| GAAGACAGTCATAAGTGCGGC
 
-
|-
 
-
|FS_77_rv|| 2013-09-11 ||Gibson-Primer rev, Amplficiation of the Backbone pSB6A1 with overlap to the RBS BBa_B0034 and the lacI-promotor, it creates an overlap to the beginning of DelH|| GCGATTTGGCGCAGGCGGCCACGGTC<br/> CATCTAGTATTTCTCCTCTTTC
 
-
|-
 
-
|FS_78_rv|| 2013-09-26 ||Gibson-Primer rev, Amplficiation of the Backbone pSB6A1 introducing the RBS BBa_B0032 and the promotor BBa_J23114  and creating an overlap to the first fragment of DelH amplified with primer DN_11|| GATTTGGCGCAGGCGGCCACGGTCCA<br/>TCTAGTACTTTCCTGTGTGACTCTAG AGCTAGCATTGTACCTAGGACTGAGCTAG<br/>CCATAAACTCTAGAAGCGGCCGCGAATTC
 
-
|-
 
-
|FS_84_fw|| 2013-09-26 ||Gibson-Primer fw, Amplficiation of the first fragment of DelH introducing the RBS BBa_B0032 and creating an overlap to primer FS_85 thereby partially introducing the promotor BBa_J23114|| GCTCAGTCCTAGGTACAATGCTAGCT<br/>CTAGAGTCACACAGGAAAGTACTAGA<br/>TGGACCGTGGCCGCCTGCG
 
-
|-
 
-
|FS_85_rv|| 2013-09-26 ||Gibson-Primer rev, Amplficiation of the Backbone pSB6A1, partially introducing the promotor BBa_B0032 with overlap to primer FS_84 and therefore the promotor BBa_J23114, it creates an overlap to the beginning of DelH|| GCGATTTGGCGCAGGCGGCCACGGTCC<br/>ATCTAGTATTTCTCCTCTTTC
 
-
|-
 
-
|FS_86_rv|| 2013-09-27 ||Gibson-Primer rev, Amplficiation of the Backbone pSB4K5 without any promotor introducing a KpnI cutting site for restriction cloning, creates an overlap to DelH and will be used for the ccdB strategy|| GGCGATTTGGCGCAGGCGGCCACGG<br/>TCCATGTACTTCGAGTCACTAAGGGCTAAC
 
-
|-
 
-
|FS_87_fw|| 2013-09-27 ||Gibson-Primer fw, Amplficiation of the Backbone pSB6A1 introducing a BamHI cutting site for restriction cloning and creating an overlap to the last fragment of DelH|| CGCTGGAGTACGCGCTGGACTGA<br/>GATCCCAGGCATCAAATAAAACG
 
-
|-
 
-
|FS_90_fw|| 2013-09-27 ||Gibson-Primer fw, Amplficiation of the ccdB cassette from the template pDonorPlasmid introducing a KpnI cutting site for restriction cloning, creates an overlap to the promotor BBa_J23114 and will be used for the ccdB strategy|| CTCAGTCCTAGGTACAATGCTAGCTCTAGA<br/>GTCACACAGGAAAGCAGTACACTGGCT<br/>GTGTATAAGGGAG
 
-
|-
 
-
|FS_93_rv|| 2013-09-27 ||Gibson-Primer rev, Amplficiation of the ccdB cassette from the template pDonorPlasmid introducing a BamHI cutting site for restriction cloning, creates an overlap to the backbone pSB6A1 and will be used for the ccdB strategy|| GTTCACCGACAAACAACAGATGA TCCGCGTGGATCCGGCTTAC
 
-
|-
 
-
|FS_94_fw|| 2013-09-27 ||Primer fw, Amplficiation of the backbone pSB6A1, will be used for the ccdB strategy|| ATCTGTTGTTTGTCGGTGAACGC
 
-
|-
 
-
| HM01:DelH_EcoRI_fw ||  -  || fw_Primer for DelH Fragment f1b|| GCATTGGAGCCTCAATGGCAAGTC
 
-
|-
 
-
|HM02:DelH_Gib1.1_rev||2013-07-09 ||Gibson-Primer DelH||  TGCTGCGCCTGCATACGGCCAAACA
 
-
|-
 
-
|HM03:DelH_Gib1.2_fw||2013-07-09 ||Gibson-Primer DelH|| AGCGGCAGGGACGACGTGGT
 
-
|-
 
-
|HM04:DelH_Gib1.2_rev||2013-07-09 ||Gibson-Primer DelH|| CATAGAGGTTGTAGAGA
 
-
|-
 
-
|HM05:DelH_Gib2.1_fw||2013-07-09 || Gibson-Primer DelH || AGAACGCCGTCTTCAGGCTCCTG
 
-
|-
 
-
|HM06:DelH_Gib2.1_rev||2013-07-09 ||Gibson-Primer DelH||  CAATGCTTTG CCGCTCGAA
 
-
|-
 
-
|HM07:DelH_Gib2.2_fw||2013-07-09 ||Gibson-Primer DelH|| TCGCCACGGCAGCTGTTCGA
 
-
|-
 
-
|HM08:DelH_Gib2_end_rev||2013-07-09 ||Gibson-Primer DelH|| TCAGTCCAGCGCGTACTCCAG
 
-
|-
 
-
|HM09:AraC_RBS_Delh_rev||2013-07-09||Gibson-Primer rev, introduces a new RBS and has the AraC-promotor and the beginning of DelH|| TTGCAAAGCGCTCGGCGATTTGGCGCAGGCG<br/>GCCACGGTCCATTTAACTTTCTCCTC<br/>TTTAATACTTTGAGCTAGCCCAA<br/>AAAAACGGTATGGAGAAACAGTAGAGAGTT
 
-
|-
 
-
|HM10:RBS_lacZ||2013-07-09 ||Gibson-Primer fw for the pSB6A1 Backbone with the end of DelH, RBS(1) and the beginning of lacZ || TGGAGTACGCGCTGGACTGA  TCTAGAG<br/>AAAGAGGAGAAA TACTAG ATGACCATGATTA
 
-
|-
 
-
|HM11:lacI_RBS(1)_DelH_rev|| 2013-07-24 ||Gibson-Primer rev, amplify the Backbone with overlap with the RBS and the lacI-promotor and it creates and overlap to the start of DelH|| TCGGCGATTTGGCGCAGGCGGCCACGGTCC<br/> ATCTAGTATTTCTCCTCTTTCTCTAGTATGTGTG
 
-
|-
 
-
|HM12:DelH_RBS(1.2)_mRFP_fw|| 2013-07-24 || Gibson-Primer fw for the pSB6A1 Backbone with the end of DelH, introducing a new RBS(new) and the beginning of mRFP|| ATTGGCGCTGGAGTACGCGCTGGACTG ATCAAAGTATTAAAGAGGA<br/>GAAAGT TAAATGGCTTCCTCCGAAGACGTTATCAAAGAG
 
-
|-
 
-
|HM13:Screen_DelH_end_fw||2013-08-16 ||New screening primer for the end of DelH together with the VR2 primer from the registry||  TTTCTGACGACCCTGCACCTGAAG
 
-
|-
 
-
|HM14:DelH_tetR_fw||2013-08-16 ||Gibson-Primer DelH-tetR: amplifies the tetracycline resistance from the pSB1T3 Backbone and creates an overlap to the end of DelH|| ATTGGCGCTGGAGTACGCGCTGGACTGA<br/>ATGAAGTTTTAAATCAATCTAAAG
 
-
|-
 
-
|HM15:tetR_stop_BB_rev||2013-08-16 ||Gibson-Primer tetR-pSB6A1: amplifies the tetracycline resistance and creates an overlap with the Terminator of the Backbone pSB6A1|| CGACTGAGCCTTTCGTTTTATTTGATGCCTGGC<br/>CTCGTGATACGCCTATTTTTATAGG
 
-
|-
 
-
|HM16:tetR_pSB6A1_fw||2013-08-16||Gibson-Primer DelH, amplifies the Backbone pSB6A1 creating an overlap with the tetracycline resistance || AAAAATAGGCGTATCACGAG  GCCAGGCA<br/>TCAAATAAAACGAAAGGCTCAG
 
-
|-
 
-
|HM17:DelH_Terminator_BB_fw|| 2013-08-16 || Gibson-Primer fw for the pSB6A1 Backbone (binding the terminator) and creating an overlap with the end of DelH|| ATTGGCGCTGGAGTACGCGCTGGACTGA<br/>AGGCATCAAATAAAACGAAAGGCTCAG
 
-
|-
 
-
| HM20:BB_HPLC_rev || 11-09-2013 || HPLC version of HM11 <br/> Gibson-Primer rev, amplify the Backbone with overlap with the <br/>RBS and the lacI-promotor and it creates and overlap to <br/>the start of DelH  || GATTTGGCGCAGGCGGCCAC<br/> GGTCCATCTAGTATTTCTCCTCTTTC
 
-
|-
 
-
| HM21:fw_lacI_BbsI_Xba || 2013-09-15 || Forward primer for cutting out mutated fragment for mutagenesis || TTTTGAAGACAA CTAGGCAATACGCAA
 
-
|-
 
-
| HM22:rev_RBS || 2013-09-15 || Reverse Primer in RBS for mutagenesis || TTTTGAAGACAA CTCTTTCTCTAGTATGTGTGAAATTG
 
-
|-
 
-
| HM23:fw_RBS || 2013-09-15 || Forward Primer in RBS for mutagenesis || TTTTGAAGACAA AGAGGAGAAATACTAGATGGACCGTGGC
 
-
|-
 
-
| HM24:rev_BbsI_MfeI || 2013-09-15 || Reverse primer for cutting out mutated fragment for mutagenesis || TTTTGAAGACAA AATTGGACAGCGCGGCATGCCGGTTG
 
-
|-
 
-
| IK01:pLF03_integr_argK_fw || 2013-06-12 || For verification of correct genomic integration of pLF03 into <i>E.coli</i> BAP1 via colony-PCR. Primer against <i>E.coli</i> BAP1 genome. || GCTGATGGAAGTGGCTGATCTGATC
 
-
|-
 
-
| IK02:pLF03_integr_pET21c_rev || 2013-06-12 || For verification of correct genomic integration of pLF03 into <i>E.coli</i> BAP1 via colony-PCR. Primer against pLF03 backbone (pET-21c). || TCCGCTCACAATTCCCCTATAGTG
 
-
|-
 
-
| IK03:pLF03_integr_argK_rev || 2013-06-18 || For verification of correct genomic integration of pLF03 into <i>E.coli</i> BAP1 via colony-PCR. Primer against <i>E.coli</i> BAP1 genome for positive control (to be used with IK01 or IK04). || GATAAATTCACTGAGCTGCCGCAG
 
-
|-
 
-
| IK04:pLF03_integr_argK_fw || 2013-06-18 || For verification of correct genomic integration of pLF03 into <i>E.coli</i> BAP1 via colony-PCR. Primer against <i>E.coli</i> BAP1 genome. Alternative forward primer in case IK01 does not work. || GCGGGAATTAATGCTGTTATGCGAAG
 
-
|-
 
-
| IK05:pLF03_integr_ygfG_fw || 2013-06-18 || For verification of correct genomic integration of pLF03 into <i>E.coli</i> BAP1 via colony-PCR. Primer against <i>E.coli</i> BAP1 genome, namely ygfG, which should be replaced by the construct. || GCGGTCATTGAGTTTAACTATGGCC
 
-
|-
 
-
| IK06:pLF03_integr_ygfG_rev || 2013-06-18 || For verification of correct genomic integration of pLF03 into <i>E.coli</i> BAP1 via colony-PCR. Primer against <i>E.coli</i> BAP1 genome, namely ygfG, which should be replaced by the construct. || AACATGGTTGAGGATGCCGACAGC
 
-
|-
 
-
| IK07:ygfG21C1 || 2013-06-25 || For insertion of methylmalonyl-CoA synthesis pathway into ''E. coli''. Extended ygfG21C1 with higher melting temperature. || TCACCGCGCACCGGCCTGCGGCAGCTCAGTGAATTTATCC<br/>AGATCTCGATCCCGCGAAATTAATAC
 
-
|-
 
-
| IK08:ygfG21C2 || 2013-06-25 || For insertion of methylmalonyl-CoA synthesis pathway into ''E. coli''. Extended ygfG21C2 with higher melting temperature. || GTTATGCTGGATAATTTCTGCCGCTTCATTGGCGGTCATC<br/>CAAAAAACCCCTCAAGACCCGTTTAG
 
-
|-
 
-
| IK24:pccB_fw || 2013-07-16 || Colony-PCR of methylmalonyl-CoA synthesis pathway || CGTATCGAGGAAGCGACGCAC
 
-
|-
 
-
| IK25:pccB_rev || 2013-07-16 || Colony-PCR of methylmalonyl-CoA synthesis pathway || GGTGATGAACATGTGGCTGGTCTG
 
-
|-
 
-
| IK26:BBa_I746200_fw || 2013-07-19 || Forward primer for permeability device || ATGAACAAGAAGATTCATTCCCTGGCCTTG
 
-
|-
 
-
| IK27:BBa_I746200-BBa_B0029_rev || 2013-07-19 || Reverse primer for permeability device with partial Gibson overhang for BBa_B0029 RBS || GAATACCAGT TCAGAAGTGGGTGTTTACGCTCATATAC
 
-
|-
 
-
| IK28:pccB-BBa_B0029-BBa_I746200_fw || 2013-07-19 || Forward primer for pccB with BBa_B0029 RBS and Gibson overhang for permeability device || ATATGAGCGTAAACACCCACTTCTGAACTGGTATTCACAC<br/>AGGAAACCTACTAG ATGTCCGAGCCGGAAGAGC
 
-
|-
 
-
| IK29:accA2-BBa_B0030_rev || 2013-07-19 || Reverse primer for accA2 with partial Gibson overhang for BBa_B0030 RBS || TAATGAAGTTG CTAGTGATTCTCGCAGATGGC
 
-
|-
 
-
| IK30:sfp-BBa_B0030-accA2_fw || 2013-07-19 || Forward primer for sfp with BBa_B0030 RBS and Gibson overhang for accA2 || TCCGGCGCCGCCATCTGCGAGAAT CACTAGCAACTTCATTAAAGAGGAG<br/>AAATACTAG ATGAAGATTTACGGAATTTATATGGAC
 
-
|-
 
-
| IK31:sfp_rev || 2013-07-19 || Reverse primer for sfp || TTATAAAAGCTCTTCGTACGAGAC
 
-
|-
 
-
| IK32:BBa_J04450-sfp_fw || 2013-07-19 || Forward primer for mRFP-containing backbones with Gibson overhang for sfp || ATCACAATGGTCTCGTACGAAGAGCTTTTATAA TACTAGAGCCAGGCATCAAATAAAACG
 
-
|-
 
-
| IK33:BBa_J04450-BBa_I746200_rev || 2013-07-19 || Reverse primer for mRFP-containing backbones with Gibson overhang for permeability device || ACAAGGCCAGGGAATGAATCTTCTTGTTCAT CTAGTATTTCTCCTCTTTCTCTAGTATG
 
-
|-
 
-
| IK34:pccB_fw || 2013-07-19 || Forward primer for pccB || ATGTCCGAGCCGGAAGAGC
 
-
|-
 
-
| IK35:BBa_J04450-pccB_rev || 2013-07-19 || Reverse primer for mRFP-containing backbones with Gibson overhang for pccB || ATGTCGGGCTGCTGCTCTTCCGGCTCGGACAT CTAGTATTTCTCCTCTTTCTCTAG
 
-
|-
 
-
| IK36:pLF03_seq_rev || 2013-07-26 || reverse primer for pLF03 for sequencing || CCGGTATCAACAGGGACACCAG
 
-
|-
 
-
| IK37:pLF03-catR_seq_rev || 2013-07-26 || reverse primer for pLF03 sequencing || CATTGAGCAACTGACTGAAATGCCTC
 
-
|-
 
-
| IK38:pIK1_fw || 2013-08-30 || Forward mutagenic primer for pIK1 || ATTCCCTGGCCTTGT T GGTCAATCTGGGGATTTATG
 
-
|-
 
-
| IK39:pIK_rev || 2013-08-30 || Reverse mutagenic primer for pIK1 || CCCAGATTGACC A ACAAGGCCAGGGAATGAATCTTC
 
-
|-
 
-
| IK40:pIK2-BBa_J23114-<br/>BBa_B0030_rev || 2013-09-06 || reverse primer for pIK2 with Gibson overhang for BBa_J23114-BBa_B0030 || TTTAATCTCTAGAGCTAGCATTGTACCTAGGACTGAGCTAGCCATAAA CTCTAGTAGAGAGCGTTCAC
 
-
|-
 
-
| IK41:BBa_I746200-BBa_B0030<br/>-BBa_J23114_fw || 2013-09-06 || forward primer for BBa_I746200 with Gibson overhang for BBa_B0030-BBa_J23114 || ATGCTAGCTCTAGAGATTAAAGAGGAGAAATACTAG ATGAACAAGAAGATTCATTCCCTG
 
-
|-
 
-
| IK42:BBa_I746200_rev || 2013-09-06 || reverse primer for BBa_I746200 || TCAGAAGTGGGTGTTTACGCTC
 
-
|-
 
-
| IK43:pIK2-BBa_I746200_fw || 2013-09-06 || forward primer for pIK2 with Gibson overhang for BBa_I746200 || TGAGCGTAAACACCCACTTCTGA TACTAGAGTCACACTGGCTC
 
-
|-
 
-
|NK_01_FS_62_screening_L_<br/>BB(without_mRFP)_rv||2013-08-02 ||Primer for screening/sequencing of pFSN construct||GTTCACCGACAAACAACAGATAAAACG
 
-
|-
 
-
| ygfG21C1 || 2013-06-03 || For insertion of methylmalonyl-CoA synthesis pathway into ''E. coli''. Primer from <bib id="pmid17959404"/>. || TCACCGCGCACCGGCCTGCGGCAGCTCAGTGAATTTATCC AGATCTCGATCCC
 
-
|-
 
-
| ygfG21C2 || 2013-06-03 || For insertion of methylmalonyl-CoA synthesis pathway into ''E. coli''. Primer from <bib id="pmid17959404"/>. || GTTATGCTGGATAATTTCTGCCGCTTCATTGGCGGTCATCC CAAAAAACCCCTCAAG
 
|-
|-
|}
|}
<br/>
<br/>
-
==Module Shuffling==
+
=Plasmids=
-
{|class="wikitable sortable"
+
-
|-
+
-
! Identifier !! Order date !! Note !! Sequence
+
-
|-
+
-
| AT01:RFC10prefix_TycA_fw|| 2013-08-12 || Fw primer for amplification of TycAdCom; introduction of RFC10 prefix || TTTT GAATTC GCGGCCGC T TCTAG ATG TTA GCA AAT CAa GCC AAT C
+
-
|-
+
-
| AT02:RFC10suffix_TycA_rv
+
-
|| 2013-08-12 || Rv primer for amplification of TycAdCom; introduction of RFC10 suffix|| TTTT CTGCAG CGGCCGC T ACTAGT A aGT TCG tTC TAC TTC TTT TTT C
+
-
|-
+
-
| AT03:RFC10prefix-TycB1_fw
+
-
|| 2013-08-12 || Fw primer for amplification of TycB1dCom; introduction of RFC10 prefix|| TTTT GAATTC GCGGCCGC T TCTAG ATG AGT GTA TTT AGC AAA GAA CAA G
+
-
|-
+
-
| AT04:RFC10suffix_TycB1_rv
+
-
|| 2013-08-12 || Rv primer for amplification of TycB1dCom; introduction of RFC10 suffix || TTTT CTGCAG CGGCCGC T ACTAGT A TTC CTC CCC aCC TTC
+
-
|-
+
-
| AT05:RFC10prefix-TycC5_fw
+
-
|| 2013-08-12 || Fw primer for amplification of TycC5; introduction of RFC10 prefix || TTTT GAATTC GCGGCCGC T TCTAG AG GCG CAT ATT GCa GAG AG
+
-
|-
+
-
| AT06:RFC10suffix_TycC5_rv
+
-
|| 2013-08-14 || Rv primer for amplification of TycC5; introduction of RFC10 suffix || TTTT CTGCAG CGGCCGC T ACTAGT A TTT GGC TGT CTC TTC GAT GAA C
+
-
|-
+
-
| AT07:RFC10prefix-TycC6_fw
+
-
|| 2013-08-12 || Fw primer for amplification of TycC6; introduction of RFC10 prefix  || TTTT GAATTC GCGGCCGC T TCTAG AG GGG AAT GTC TTC TCG ATC
+
-
|-
+
-
| AT08:RFC10suffix_TycC6_rv
+
-
|| 2013-08-14 || Rv primer for amplification of TycC6; introduction of RFC10 suffix || TTTT CTGCAG CGGCCGC T ACTAGT A TTA TTT CAG GAT aAA CAG TTC TTG
+
-
|-
+
-
| AT09:R10_B1_fw_longer
+
-
|| 2013-08-18 || Fw primer (longer) for amplification of TycB1dCom; introduction of RFC10 prefix  || TTTT GAA TTC GCG GCC GCT TCT AG    ATG AGT GTA TTT AGC AAA GAA CAA GTT C
+
-
|-
+
-
| AT10:R10_B1_rv_longer
+
-
|| 2013-08-18 || Rv primer (longer for amplification of TycB1dCom; introduction of RFC10 suffix || TTTT CTG CAG CGG CCG CTA CTA GTA ATA CGC aCT TTC CTC CCC GCC
+
-
|-
+
-
| AT11:R10_C5_fw_rpos
+
-
|| 2013-08-18 || Fw primer (repositioned) for amplification of TycC6; introduction of RFC10 prefix|| TTTT GAATTC GCGGCCGC T TCTAG AG GAGCAGTTCGAGACGATCCAGCC
+
-
|-
+
-
| AT101|| 2013-09-05|| forward screening primer TycAdCom || GGACATCATCGAACAGGC
+
-
|-
+
-
| AT102 || 2013-09-05|| reverse screening primer TycAdCom || GACATAGGCAAGATCCGTAG
+
-
|-
+
-
| AT103|| 2013-09-05|| forward screening primer TycAdCom || GCAAGGACAAGTAGATGGC
+
-
|-
+
-
| AT104 || 2013-09-05|| reverse screening primer TycAdCom || GTACAGCCATACAGTCGC
+
-
|-
+
-
| AT105 || 2013-09-05|| forward screening primer TycC5 || GTCACCATGCTGAGAACG
+
-
|-
+
-
| AT106 || 2013-09-05|| reverse screening primer TycC5 || CAGCAAAGCGGTCATAATC
+
-
|-
+
-
| AT107 || 2013-09-05|| forward screening primer TycC6 || AAGCTACGCTGTTGATTGC
+
-
|-
+
-
| AT108 ||  2013-09-05|| reverse screening primer TycC6 || AGCACGTAACGATCCTC
+
-
|-
+
-
| IK09:TycA_A1_fw || 2013-07-08 || Colony-PCR of ''Brevibacillus parabrevis'': TycA A domain || ATGTTAGCAAATCAAGCCAATCTC
+
-
|-
+
-
| IK10:TycA_A1_rev || 2013-07-08 || Colony-PCR of ''Brevibacillus parabrevis'': TycA A domain || TTGGTTTGCTGTAAGATCAGGCTC
+
-
|-
+
-
| IK11:TycB_A1_fw || 2013-07-08 || Colony-PCR of ''Brevibacillus parabrevis'': TycB A1 domain || AATTCGGGAGTCGAGCTTTGTCAG
+
-
|-
+
-
| IK12:TycC6_rev || 2013-07-19 || reverse primer for tyrocidine TycC6 module || TTATTTCAGGATGAACAGTTCTTGCAGG
+
-
|-
+
-
| IK13:IK13:TycB1-dCom-dC_fw || 2013-07-19 || forward primer for tyrocidine TycB1 module without Com and C domain || GATTGCGTGGCAAACAATTCGGGAGTC
+
-
|-
+
-
| IK14:TycB1-TycC6_rev || 2013-07-19 || reverse primer for tyrocidine TycB1 module with Gibson overhang for TycC6 || CAGGCTCGATCGAGAAGACATTCCC TTCCTCCCCGCCTTCCACATACGC
+
-
|-
+
-
| IK15:TycC6-TycB1_fw || 2013-07-19 || forward primer for tyrocidine TycC6 module with Gibson overhang for TycB1 || GCGTATGTGGAAGGAGGGGAGGAA GGGAATGTCTTCTCGATCGAGCCTG
+
-
|-
+
-
| IK16:TycA_fw || 2013-07-19 || forward primer for tyrocidine TycA module || ATGTTAGCAAATCAGGCCAATCTCATC
+
-
|-
+
-
| IK17:TycA-dCom-TycC5_rev || 2013-07-19 || reverse primer for tyrocidine TycA module without Com domain with Gibson overhang for TycC5 || GAATGCGCTCTCGGCAATATGGGC TGTTCGCTCTACTTCTTTTTTCTCGG
+
-
|-
+
-
| IK18:TycC5-TycA-dCom_fw || 2013-07-19 || forward primer for tyrocidine TycC5 module with Gibson overhang for TycAdCom || CCGAGAAAAAAGAAGTAGAGCGAACA GCCCATATTGCCGAGAGCGCATTC
+
-
|-
+
-
| IK19:TycB1-TycC5_rev || 2013-07-19 || reverse primer for tyrocidine TycB1 module with Gibson overhang for TycC5 || GAATGCGCTCTCGGCAATATGGGC TTCCTCCCCGCCTTCCACATACGC
+
-
|-
+
-
| IK20:TycC5-TycB1_fw || 2013-07-19 || forward primer for tyrocidine TycC5 module with Gibson overhang for TycB1 || GCGTATGTGGAAGGAGGGGAGGAA GCCCATATTGCCGAGAGCGCATTC
+
-
|-
+
-
| IK21:pSB4K5-TycB1dComdC_rev || 2013-07-19 || reverse primer for mRFP-carrying backbones with Gibson overhang for tyrocidine TycA module (nomenclature wrong) || GTCGATGAGATTGGCCTGATTTGCTAACAT CTAGTATTTCTCCTCTTTCTCTAGTATGTG
+
-
|-
+
-
| IK22::pSB4K5-TycC6_fw || 2013-07-19 || forward primer for mRFP-carrying backbones with Gibson overhang for tyrocidine TycC6 module || AACATCCTGCAAGAACTGTTCATCCTGAAA TAATAACGCTGATAGTGCTAGTGTAGATC
+
-
|-
+
-
| IK23:pSB4K5-TycA_rev || 2013-07-19 || reverse primer for mRFP-carrying backbones with Gibson overhang for tyrocidine TycB1 module without Com and C domains + start codon (nomenklature wrong) || GACTCCCGAATTGTTTGCCACGCAATCCAT CTAGTATTTCTCCTCTTTCTCTAGTATGTG
+
-
|-
+
-
| PW01:TycB_A1_rev || 2013-07-08 || Colony-PCR of ''Brevibacillus parabrevis'': TycB A1 domain || CTTGGCACTTCCTTCAGGCTTC
+
-
|-
+
-
| PW02:TycB_E1_fw || 2013-07-08 || Colony-PCR of ''Brevibacillus parabrevis'': TycB E domain || CGAGAGAGCGAGCAAGGTG
+
-
|-
+
-
| PW03:TycB_E1_rev || 2013-07-08 || Colony-PCR of ''Brevibacillus parabrevis'': TycB E domain || GTACTCGCCTTCTTCTTTTGC
+
-
|-
+
-
| PW04:pSB1C3-TycC5ΔC_rev || 2013-07-19 || Integration of Tetrapeptide NRPS from ''Brevibacillus parabrevis'' in backbone; Gibson primer for Tetrapeptide I & II || TGTTTTGGTTGCGAGGAAGCTGTGCAGCAT CTAGTATTTCTCCTCTTTCTCTAGTATGTG
+
-
|-
+
-
| PW05:TycC5ΔC_fwd || 2013-07-19 || Amplification of TycC5-Module from ''Brevibacillus parabrevis''; Gibson primer for Tetrapeptide I & II || ATGCTGCACAGCTTCCTCGCAACCAAAACAGCC
+
-
|-
+
-
| PW06:TycC5ΔC-TycB1ΔCom_rev || 2013-07-19 || Amplification of TycC5-Module from ''Brevibacillus parabrevis''; Gibson primer for Tetrapeptide I & II || GTGAAACAGCATCCCCTCTTGCATCGG AGGCTCGATCGAGAAGACATTCCCTTTG
+
-
|-
+
-
| PW07:TycC5ΔC-TycB1ΔCom_fwd || 2013-07-19 || Amplification of TycB1+C(TycB2)-Module from ''Brevibacillus parabrevis''; Gibson primer for Tetrapeptide I & II || CAAAGGGAATGTCTTCTCGATCGAGCCT CCGATGCAAGAGGGGATGCTGTTTCAC
+
-
|-
+
-
| PW08:C(TycB2)-TycAΔCom_rev || 2013-07-19 || Amplification of TycB1+C(TycB2)-Module from ''Brevibacillus parabrevis''; Gibson primer for Tetrapeptide I & II || GTTGTCGATGAGATTGGCCTGATTTGCTAACAT GATTTGCGCCAGCTCCTGCTCCGTGTT
+
-
|-
+
-
| PW09:C(TycB2)-TycAΔCom_fwd || 2013-07-19 || Amplification of TycA-Module from ''Brevibacillus parabrevis''; Gibson primer for Tetrapeptide I & II || AACACGGAGCAGGAGCTGGCGCAAATC ATGTTAGCAAATCAGGCCAATCTCATCGACAAC
+
-
|-
+
-
| PW10:TycAΔCom-TycC6_rev || 2013-07-19 || Amplification of TycAΔCom-Module from ''Brevibacillus parabrevis''; Gibson primer for Tetrapeptide I || CTTTGGCTGTCTCTTCGATGAACGC TCGCTCTACTTCTTTTTTCTCGGTGCAATG
+
-
|-
+
-
| PW11:TycAΔCom-TycC6_fwd || 2013-07-19 || Amplification of TycC6-Module from ''Brevibacillus parabrevis''; Gibson primer for Tetrapeptide I || CATTGCACCGAGAAAAAAGAAGTAGAGCGA GCGTTCATCGAAGAGACAGCCAAAG
+
-
|-
+
-
| PW12:TycAΔE-TycC6_rev || 2013-07-19 || Amplification of TycAΔE-Module from ''Brevibacillus parabrevis''; Gibson primer for Tetrapeptide II; one mismatch C->t || CTTTTGCACAGGCTCGATCGAGAAGAC GCTtTTGACAAAAAGAGCAACCTG
+
-
|-
+
-
| PW13:TycAΔE-TycC6_fwd || 2013-07-19 || Amplification of TycC6-Module from ''Brevibacillus parabrevis''; Gibson primer for Tetrapeptide II; one mismatch G->a || CAGGTTGCTCTTTTTGTCAAaAGC GTCTTCTCGATCGAGCCTGTGCAAAAG
+
-
|-
+
-
| PW14:C(TycC2)-indC_rev || 2013-08-16 || Amplification of C-domain from TycC2 from ''Brevibacillus parabrevis''; Gibson overhang to IndC; for construct 1, 2 & 3 || ACATTGTGTAATATTATTTTCTAACAT CGTTTTGCTGCTGGCAGGCTG
+
-
|-
+
-
| PW15:C(TycC2)-indC_fwd || 2013-08-16 || Amplification of indC from ''Photorhabdus luminescens''; Gibson overhang to C-domain from TycC2; for construct 1, 2 & 3 || CAGCCTGCCAGCAGCAAAACG ATGTTAGAAAATAATATTACACAATGT
+
-
|-
+
-
| PW16:indC_rev || 2013-08-16 || Amplification of indC from ''Photorhabdus luminescens''; no Gibson overhang; for construct 1, 2 & 3 || TTAGATTATTTTCTCAATCTCAGCAACACCTTC
+
-
|-
+
-
| PW17:TycAdE-C(TycC2)_rev || 2013-08-16 || Amplification of TycAdE from ''Brevibacillus parabrevis''; Gibson overhang to C-domain from TycC2; for construct 1 || CGAAAGGAAGCGGGCCAGCTC AGCAACCTGCTCGATCGTCGGGTA
+
-
|-
+
-
| PW18:TycAdE-C(TycC2)_fwd || 2013-08-16 || Amplification of C-domain from TycC2-module from ''Brevibacillus parabrevis''; Gibson overhang to TycAdE; for construct 1 || TACCCGACGATCGAGCAGGTTGCT GAGCTGGCCCGCTTCCTTTCG
+
-
|-
+
-
| PW19:TycC4dC_fwd || 2013-08-16 || Amplification of TycC4-module from ''Brevibacillus parabrevis'' without the C-domain; no Gibson overhang, ATG added; for construct 3 || ATGTATCCGCGCGATCTGACGATTC
+
-
|-
+
-
| PW20:TycC4dC-C(TycC2)_rev || 2013-08-16 || Amplification of TycC4-module from ''Brevibacillus parabrevis'' without the C-domain; Gibson overhang to C-domain form TycC2; for construct 3 || GGTGTACTCGGTTTTTTCCGA AATATGCGCAGCCAACTCATG
+
-
|-
+
-
| PW21:TycC4dC-C(TycC2)_fwd || 2013-08-16 || Amplification of C-domain from TycC2-module from ''Brevibacillus parabrevis''; Gibson overhang to TycC4dC; for construct 3 || CATGAGTTGGCTGCGCATATT TCGGAAAAAACCGAGTACACC
+
-
|-
+
-
| PW22:pSB1C3-TycC4dC_rev || 2013-08-16 || Insertion of construct 3 into pSB1C3-backbone, amplification of pSB1C3; Gibson overhang to TycC4dC; for construct 3 || GAATCGTCAGATCGCGCGGATACAT CTAGTATTTCTCCTCTTTCTCTAGTATGTG
+
-
|-
+
-
| PW23:indC-pSB1C3_fwd || 2013-08-16 || Insertion of either fragments into pSB1C3-backbone, amplification of pSB1C3; Gibson overhang to indC from ''Photorhabdus luminescens''; for constructs 1, 2 & 3 || GGTGTTGCTGAGATTGAGAAAATAATCTAA TAATAACGCTGATAGTGCTAGTGTAGATC
+
-
|-
+
-
| PW24:TycC1dC_fwd || 2013-08-16 || Amplification of the TycC1-module from ''Brevibacillus parabrevis'' without the C-domain; no Gibson overhang, ATG added; for construct 2 || ATGCAGACGAACAAACAACAGACG
+
-
|-
+
-
| PW25:pSB1C3-TycC1dC || 2013-08-16 || Insertion of construct 2 in pSB1C3-backbone, amplification of pSB1C3; Gibson overhang to TycC1-module without C-domain; for construct 2 || CGTCTGTTGTTTGTTCGTCTGCAT CTAGTATTTCTCCTCTTTCTCTAGTATGTG
+
-
|-
+
-
| PW26:TycC5dC_fwd || 2013-09-04 || Amplification of TycC5-module without C-domain from ''Brevibacillus parabrevis''; no Gibson overhang, ATG added; for constructs A & B || ATGCTGCACAGCTTCCTCGCAACC
+
-
|-
+
-
| PW27:TycC5dC-C(TycC4)_rev || 2013-09-04 || Amplification of TycC5-module without C-domain from ''Brevibacillus parabrevis''; Gibson overhang to C-domain from TycC4; for constructs A & B || CACATACGTCTCTTTTCCGCTCGT TTCGATGAACGCCGCCAGTTC
+
-
|-
+
-
| PW28:TycC5dC-C(TycC4)_fwd || 2013-09-04 || Amplification of C-domain from TycC4-module from ''Brevibacillus parabrevis''; Gibson overhang to TycC5-module without C-domain; for constructs A & B || GAACTGGCGGCGTTCATCGAA ACGAGCGGAAAAGAGACGTATGTG
+
-
|-
+
-
| PW29:C(TycC4)-TycC4dC_rev || 2013-09-04 || Amplification of C-domain from TycC4-module from ''Brevibacillus parabrevis''; Gibson overhang to TycC4-module without C-domain; for constructs A, B, C, E & G || GAATCGTCAGATCGCGCGGATA GGCAAACGTGTTGTTGAAATC
+
-
|-
+
-
| PW30:C(TycC4)-TycC4dC_fwd || 2013-09-04 || Amplification of TycC4-module without C-domain from ''Brevibacillus parabrevis''; Gibson overhang to C-domain from TycC4-module; for constructs A, B, C, E & G || GATTTCAACAACACGTTTGCC TATCCGCGCGATCTGACGATTC
+
-
|-
+
-
| PW31:TycC4-C(TycC4)_rev || 2013-09-04 || Amplification of TycC4-module from ''Brevibacillus parabrevis''; Gibson overhang to C-domain from TycC4-module; for constructs B & C || CACATACGTCTCTTTTCCGCTCGT GGCAATATGCGCAGCCAACTCATG
+
-
|-
+
-
| PW32:TycC4-C(TycC4)_fwd || 2013-09-04 || Amplification of C-domain from TycC4-module from ''Brevibacillus parabrevis''; Gibson overhang to TycC4-module; for constructs B & C || CATGAGTTGGCTGCGCATATTGCC ACGAGCGGAAAAGAGACGTATGTG
+
-
|-
+
-
| PW33:TycC6dTE-C(TycC2)_rev || 2013-09-04 || Amplification of TycC6-module without the TE-domain from ''Brevibacillus parabrevis''; Gibson overhang to C-domain from TycC2-module; for constructs D & F || GGTGTACTCGGTTTTTTCCGA CGTGATGAAATCGGCCACCTTTTC
+
-
|-
+
-
| PW34:TycC6dTE-C(TycC2)_fwd || 2013-09-04 || Amplification of C-domain from TycC2-module from ''Brevibacillus parabrevis''; Gibson overhang to TycC6-module without TE-domain; for constructs D & F || GAAAAGGTGGCCGATTTCATCACG TCGGAAAAAACCGAGTACACC
+
-
|-
+
-
| PW35:TycC6dTE-C(TycC4)_rev || 2013-09-04 || Amplification of TycC6-module without the TE-domain from ''Brevibacillus parabrevis''; Gibson overhang to C-domain from TycC4-module; for constructs E & G || CACATACGTCTCTTTTCCGCTCGT CGTGATGAAATCGGCCACCTTTTC
+
-
|-
+
-
| PW36:TycC6dTE-C(TycC4)_fwd || 2013-09-04 || Amplification of C-domain from TycC4-module from ''Brevibacillus parabrevis''; Gibson overhang to TycC6-module without TE-domain; for constructs E & G || GAAAAGGTGGCCGATTTCATCACG ACGAGCGGAAAAGAGACGTATGTG
+
-
|-
+
-
| PW37:pSB1C3-TycC5dC_rev || 2013-09-04 || Insertion of constructs A & B in pSB1C3-backbone, amplification of pSB1C3; Gibson overhang to TycC5-module without C-domain; for constructs A & B || GGTTGCGAGGAAGCTGTGCAGCAT CTAGTATTTCTCCTCTTTCTCTAGTATGTG
+
-
|}
+
-
<br/>
+
-
==Linker Variation==
+
-
{|class="wikitable sortable"
+
-
|-
+
-
! Identifier !! Order date !! Note !! Sequence
+
-
|-
+
-
|AR01|| 2013-09-17 || sTycC4dC_fw || ATG AAGCACTTGCTGCCGCTCGTC
+
-
|-
+
-
|AR02|| 2013-09-17 ||c(TycC2)s_rev|| TTGTGTAATATTATTTTCTAACAT CGCTACCTGTTGCAACCGCTC
+
-
|-
+
-
|AR03|| 2013-09-17 ||s-Ind_fw|| GAGCGGTTGCAACAGGTAGCG ATGTTAGAAAATAATATTACAATGT
+
-
|-
+
-
|AR04|| 2013-09-17 ||c(TycC2)l_rev_l||TTGTGTAATATTATTTTCTAACAT AAGCGACTGCTGGTTCGCG
+
-
|-
+
-
|AR05|| 2013-09-17 ||l-Ind_fw||AACGCGAACCAGCAGTCGCTT ATGTTAGAAAATAATATTACACAATGT
+
-
|-
+
-
|AR06|| 2013-09-17 ||lTycC4dC_fw||ATG CGCAACTATCCGGTCGAGACG
+
-
|-
+
-
|AR07|| 2013-09-17 ||TycC4dCom_short-BB_rev||GACGAGCGGCAGCAAGTGCTTCAT CTAGTATTTCTCCTCTTTCTCTAGTATGTG
+
-
|-
+
-
|AR08|| 2013-09-17 ||TycC4dCom_long-BB_rev||CGTCTCGACCGGATAGTTGCGCAT CTAGTATTTCTCCTCTTTCTCTAGTATGTG
+
-
|-
+
-
| PW19:TycC4dC_fwd || 2013-08-16 || Amplification of TycC4-module from ''Brevibacillus parabrevis'' without the C-domain; no Gibson overhang, ATG added; for construct 3 || ATGTATCCGCGCGATCTGACGATTC
+
-
|-
+
-
| PW20:TycC4dC-C(TycC2)_rev || 2013-08-16 || Amplification of TycC4-module from ''Brevibacillus parabrevis'' without the C-domain; Gibson overhang to C-domain form TycC2; for construct 3 || GGTGTACTCGGTTTTTTCCGA AATATGCGCAGCCAACTCATG
+
-
|-
+
-
| PW21:TycC4dC-C(TycC2)_fwd || 2013-08-16 || Amplification of C-domain from TycC2-module from ''Brevibacillus parabrevis''; Gibson overhang to TycC4dC; for construct 3 || CATGAGTTGGCTGCGCATATT TCGGAAAAAACCGAGTACACC
+
-
|-
+
-
| PW14:C(TycC2)-indC_rev || 2013-08-16 || Amplification of C-domain from TycC2 from ''Brevibacillus parabrevis''; Gibson overhang to IndC; for construct 1, 2 & 3 || ACATTGTGTAATATTATTTTCTAACAT CGTTTTGCTGCTGGCAGGCTG
+
-
|-
+
-
|}
+
-
<br/>
+
-
==Domain Shuffling and PPTases==
+
-
{|class="wikitable sortable"
+
-
|-
+
-
! Identifier !! Order date !! Note !! Sequence
+
-
|-
+
-
| KH1_ccdb_fw || 2013-07-23 || amplifing ccdb with RBS from pDONR || TGGTGCCATTACATACAGATACT GAGAACAGGGGCTGGTGAAATGC
+
-
|-
+
-
| KH2_ccdb_rv || 2013-07-23 || amplifing ccdb with RBS from pDONR || AGAGTCTGTCTGTTCAATCCACTT TTATATTCCCCAGAACATCAGGTTAATGGCG
+
-
|-
+
-
| KH3_indC_backbone-withoutT_fw || 2013-07-23 || linearization of vector with partial indigoidine synthase || AAGTGGATTGAACAGACAGACTCTAAAAC
+
-
|-
+
-
| KH4_indC_backbone-withoutT_rv || 2013-07-23 || linearization of vector with partial indigoidine synthase || AGTATCTGTATGTAATGGCACCAATAGACGC
+
-
|-
+
-
| KH5_indC_T_fw || 2013-07-23 || introducing T domain of indC || TGGTGCCATTACATACAGATACT GAAATAAGGCTTGGAAAAATTTGGATGGAAGT
+
-
|-
+
-
| KH6_indC_T_rv || 2013-07-23 || introducing T domain of indC || AGAGTCTGTCTGTTCAATCCACTT AGCCAATTCTGCTATATTAGGAGATTGA
+
-
|-
+
-
| KH7_bpsA_T_fw || 2013-07-23 || introducing T domain of bpsA || TGGTGCCATTACATACAGATACT GAGAAGGAAATCGCAGCCGTGTGGG
+
-
|-
+
-
| KH8_bpsA_T_rv || 2013-07-23 || introducing T domain of bpsA || AGAGTCTGTCTGTTCAATCCACTT GGCCAGCTTTTCAATTGTTGGTGACTCC
+
-
|-
+
-
| KH9_ccdB-Big_fw || 2013-08-06 || ccdB_Big amplified from pDONOR (Dominik) for indC(ccdB) construct || TGCCATTACATACAGATACT ACTGGCTGTGTATAAGGGAGCCTGAC
+
-
|-
+
-
| KH10_ccdB-Big_rv || 2013-08-06 || ccdB_Big amplified from pDONOR (Dominik) for indC(ccdB) construct || AGAGTCTGTCTGTTCAATCCACTT CGCGTGGATCCGGCTTAC
+
-
|-
+
-
| KH11_plu2642-T_fw || 2013-08-14 || T domain exchange plu2642 || TGGTGCCATTACATACAGATACT GAAACACAGATCGTAAAGATATGG
+
-
|-
+
-
| KH12_plu2642-T_rv || 2013-08-14 || T domain exchange plu2642 || AGAGTCTGTCTGTTCAATCCACTT TGCCAATTGTTTAACCGTTG
+
-
|-
+
-
| KH13_BBa-entD_fw || 2013-08-09 || submitting endD to registry || GCAT GAATTCGCGGCCGCTTCTAG ATGAAAACTACGCATACCTCCCTC
+
-
|-
+
-
| KH14_BBa-entD_rv || 2013-08-09 || submitting endD to registry || GCAT CTGCAGCGGCCGCTACTAGTA TCATTAATCGTGTTGGCACAGCG
+
-
|-
+
-
| KH15_BBa-sfp_Bsub_fw || 2013-08-09 || submitting sfp_Bsub to registry || GCAT GAATTCGCGGCCGCTTCTAG ATGAAGATTTACGGAATTTATATGGAC
+
-
|-
+
-
| KH16_BBa-sfp_Bsub_rv || 2013-08-09 || submitting sfp_Bsub to registry || GCAT CTGCAGCGGCCGCTACTAGTA TCATTATAAAAGCTCTTCGTACGAGAC
+
-
|-
+
-
| KH17_BBa-svp_Svert_fw || 2013-08-09 || submitting svp_Svert to registry || GCAT GAATTCGCGGCCGCTTCTAG A TGATCGCCGCCCTCCTGCCCTCC
+
-
|-
+
-
| KH18_BBa-svp_Svert_rv || 2013-08-09 || submitting svp_Svert to registry || GCAT CTGCAGCGGCCGCTACTAGTA AGCTAGCTCATTACGGGAC
+
-
|-
+
-
| KH19_BBa-delC_fw || 2013-08-09 || submitting delC to registry || GCAT GAATTCGCGGCCGCTTCTAG ATGCAGCTCGTGTCCGTGCG
+
-
|-
+
-
| KH20_BBa-delC_rv || 2013-08-09 || submitting delC to registry || GCAT CTGCAGCGGCCGCTACTAGTA TTA TCATGTCGATTCCTTGGTGC
+
-
|-
+
-
| KH21_BBa-indC_fw || 2013-08-09 || submitting indC to registry || GCAT GAATTCGCGGCCGCTTCTAG ATGTTAGAAAATAATATTACACAATG
+
-
|-
+
-
| KH22_BBa-indC_rv || 2013-08-09 || submitting indC to registry || GCAT CTGCAGCGGCCGCTACTAGTA TTAGATTATTTTCTCAATCTCAG
+
-
|-
+
-
| NI01:bpsA_AOxA_PstI_fw|| 2013-06-04 || for amplifying bpsA (AOxA domain) || GAGGAGAAATACTAGATGACACTGCAGGAAACAAGCGTGC
+
-
|-
+
-
| NI02:bpsA_AOxA_rv|| 2013-06-04 || for amplifying bpsA (AOxA domain)  || GAAGGGCCGTTCCACCAGCTCAGCGTTGACCTGGTCAGAAGCG
+
-
|-
+
-
| NI03:bpsA_T_fw|| 2013-06-04 || for amplifying bpsA (T domain) || GACCAGGTCAACGCTGAGCTGGTGGAACGGCCCTTCGTCG
+
-
|-
+
-
| NI04:bpsA_T_rv|| 2013-06-04 || for amplifying bpsA (T domain) || GACGAAGCGACTAGACTCCTGAGCCACTTCTCTCTCCAGCCG
+
-
|-
+
-
| NI05:bpsA_TE_fw|| 2013-06-04 || for amplifying bpsA (TE domain) with RBS 1 || GAGAGAGAAGTGGCTCAGGAGTCTAGTCGCTTCGTCCGA
+
-
|-
+
-
| NI06:bpsA_TE_Stop_XbaI_rbs_rv|| 2013-06-04 || for amplifying bpsA (TE domain) with RBS 1 || TGGCAGCAGAGCAGCGATCATCTAGTATTT CTCCTCTTTCCTCTAGATCATCATTCCC CCAGCAGGTATCTAAT
+
-
|-
+
-
| NI07:svp_fw|| 2013-06-04 || for amplifying svp || GAGGAGAAATACTAGATGATCGCTGCTCTGCTGCCAAGTTGG
+
-
|-
+
-
| NI08:svp_rv|| 2013-06-04 || for amplifying svp || GCACTATCAGCGTTATTATGGCACGGCAGTCCTATCGTCG
+
-
|-
+
-
| NI09:pSB1C3_fw|| 2013-06-04 || for linearizing, amplifying pSB1C3 || GATAGGACTGCCGTGCCATAATAA CGCTGATAGTGCTAGTGTAGATCGC
+
-
|-
+
-
| NI10:pSB1C3_PstI_rv|| 2013-06-04 || for linearizing, amplifying pSB1C3 || GCTTGTTTCCTGCAGTGTCATCTAG TATTTCTCCTCTTTCTCTAGTATGTG
+
-
|-
+
-
| NI11:bpsA_rvN|| 2013-07-15 || for amplifying bpsA (TE domain) with RBS 2 || TGGCAGCAGAGCAGCGATCATTATTTAGGT<br/>TTCCTGTGTGAATCATCATTCCCCCAGCAGGTATCTAAT
+
-
|-
+
-
| NI12:svp_fwN|| 2013-07-15 || for amplifying svp with RBS 2  || CAGGAAACCTAAATAATGATC GCTGCTCTGCTGCCAAGTTGG
+
-
|-
+
-
| NI13_ccdb_fw || 2013-07-23 || amplifing ccdb with RBS from pDONR || CGTCGCACCTAGGACCGAAACA GAGAACAGGGGCTGGTGAAATGC
+
-
|-
+
-
| NI14_ccdb_rv || 2013-07-23 || amplifing ccdb with RBS from pDONR || GCCACTTCTCTCTCCAGCCGTCG TTATATTCCCCAGAACATCAGGTTAATGGCG
+
-
|-
+
-
| NI15_bpsA_backbone-withoutT_fw || 2013-07-23 || linearization of vector with partial indigoidine synthase || CGACGGCTGGAGAGAGAAGTGGC
+
-
|-
+
-
| NI16_bpsA_backbone-withoutT_rv || 2013-07-23 || linearization of vector with partial indigoidine synthase || TC TGTTTCGGTCCTAGGTGCGACG
+
-
|-
+
-
| NI17_indC_T_fw || 2013-07-23 || introducing T domain of indC || CGTCGCACCTAGGACCGAAACA GAAATAAGGCTTGGAAAAATTTGGATGGAAGT
+
-
|-
+
-
| NI18_indC_T_rv || 2013-07-23 || introducing T domain of indC || GCCACTTCTCTCTCCAGCCGTCG AGCCAATTCTGCTATATTAGGAGATTGA
+
-
|-
+
-
| NI19_bpsA_T_fw || 2013-07-23 || introducing T domain of bpsA || CGTCGCACCTAGGACCGAAACA GAGAAGGAAATCGCAGCCGTGTGGG
+
-
|-
+
-
| NI20_bpsA_T_rv || 2013-07-23 || introducing T domain of bpsA || GCCACTTCTCTCTCCAGCCGTCG GGCCAGCTTTTCAATTGTTGGTGACTCC
+
-
|-
+
-
| NI19:ngrA_fw || || || GAATTCGCGGCCGCTTCTAG ATGGAACAGACAGTTATACACACC
+
-
|-
+
-
| NI20:ngrA_rv || || || CTGCAGCGGCCGCTACTAGTA TTATTGAATAATTAGCGTTAATATTTCCTG
+
-
|-
+
-
|RB05:RBS1-BpsA_fw||2013-07-01||Gibson assembly of pSB1C3 bpsA svp||AAAGAGGAGAAA TACTAG ATGACACTTCAGGAAACCAGCG
+
-
|-
+
-
|RB06:BpsA-RBS2_rv||2013-07-01||Gibson assembly of pSB1C3 bpsA svp||CTTTCCTGTGTGA AAGCTT TCA TCACTCCCCGAGCAGATATCG
+
-
|-
+
-
|RB07:RBS2-svp_fw||2013-07-01||Gibson assembly of pSB1C3 bpsA svp||CTTTCACACAGGAAAG TAAATA ATGGCTGCTCTTCTTCCTAGTTGGGC
+
-
|-
+
-
|RB08:svp-pSB1C3_rv||2013-07-01||Gibson assembly of pSB1C3 bpsA svp||CTATCAGCGTTATTA AAGCTT TCATCA TGGCACGGCAGTCCTATCG
+
-
|-
+
-
|RB09:pSB1C3_fw||2013-07-01||Gibson assembly of pSB1C3 bpsA svp||AAGCTT TAATAACGCTGATAGTGCTAGTGTAGATCGC
+
-
|-
+
-
|RB10:pSB1C3-RBS1_rv||2013-07-01||Gibson assembly of pSB1C3 bpsA svp||CTAGTA TTTCTCCTCTTT CTCTAGTATGTGTG
+
-
|-
+
-
|RB11:BpsA_Taka_fw||2013-07-08||colony PCR bpsA Takahashi||CAT ATGACTCTTCAGGAGACCAGCGTGCTC
+
-
|-
+
-
|RB12:BpsA_Taka_rv||2013-07-08||colony PCR bpsA Takahashi||AAG CTTCTCGCCGAGCAGGTAGCGGATGTG
+
-
|-
+
-
|RB13:svp_Taka_fw||2013-07-08||colony PCR svp Takahashi||CAT ATGATCGCCGCCCTCCTGCCCTCCTG
+
-
|-
+
-
|RB14:svp_Taka_rv||2013-07-08||colony PCR svp Takahashi||CTCGAGCGGGACGGCGGTCCGGTCGTCCGC
+
-
|-
+
-
|RB15:svp_Sanch_fw||2013-07-08||PCR svp Sanchez||TATA ATGCATGCTCGCCGCCCTCCCC
+
-
|-
+
-
|RB16:svp_Sanch_rv||2013-07-08||PCR svp Sanchez||TTAAGATCTCGGGACGGCGGTCCGGTC
+
-
|-
+
-
|RB17:sfp_fw||2013-07-08||sfp extraction from <i>E.coli</i> BAP1||ATGAAGATTTACGGAATTTATATGG
+
-
|-
+
-
|RB18:sfp_rv||2013-07-08||sfp extraction from <i>E.coli</i> BAP1||TTATAAAAGCTCTTCGTACGAGACC
+
-
|-
+
-
|RB19:entD_fw||2013-07-08||entD extraction from <i>E.coli</i> Lambalot||TAAATA ATGGTCGATATGAAAACTACGC
+
-
|-
+
-
|RB20:entD_rv||2013-07-08||entD extraction from <i>E.coli</i> Lambalot||AAGCTT ATTAATCGTGTTGGCACAGCG
+
-
|-
+
-
| RB21:pSB1C3_fw || 2013-07-15 || indigoidine exchangeable construct || TAATGA GCTAGC TAATAACGCTGATAGTGCTAGTG
+
-
|-
+
-
| RB22:pSB1C3_rv || 2013-07-15 || indigoidine exchangeable construct || CAT GGTACC TTTCTCCTCTTT CTCTAGTATGTGTG
+
-
|-
+
-
| RB23:bpsA-fus_fw || 2013-07-15 || indigoidine exchangeable construct || CACACATACTAGAG AAAGAGGAGAAA GGTACC ATGACTAGTACACTGCAGG
+
-
|-
+
-
| RB24:bpsA-fus_rv || 2013-07-15 || indigoidine exchangeable construct || CAT GGATCC GGTTTCCTGTGTGAA TCATTA TTCCCCCAGCAGGTATCTAATATG
+
-
|-
+
-
| RB25:svp-fus_fw || 2013-07-15 || indigoidine exchangeable construct || CACAGGAAACC GGATCC ATGACTAGTATCGCTGCTCTGCTG
+
-
|-
+
-
| RB26:svp-fus_rv || 2013-07-15 || indigoidine exchangeable construct || CAGCGTTATTA GCTAGC TCATTA TGGCACGGCAGTCCTATC
+
-
|-
+
-
| RB27:indC-Plum_fw || 2013-07-15 || indigoidine exchangeable construct || CATACTAGAG AAAGAGGAGAAA GGTACC ATGTTAGAAAATAATATTACACAATG
+
-
|-
+
-
| RB28:indC-Plum_rv || 2013-07-15 || indigoidine exchangeable construct || CAT GGATCC GGTTTCCTGTGTGAA TTA TTAGATTATTTTCTCAATCTCAG
+
-
|-
+
-
| RB29:svp-Svert_fw || 2013-07-15 || indigoidine exchangeable construct || CACAGGAAACC GGATCC A TGATCGCCGCCCTC
+
-
|-
+
-
| RB30:svp-Svert_rv || 2013-07-15 || indigoidine exchangeable construct || CAGCGTTATTA GCTAGC TCA TTACGGGACGGCGGTC
+
-
|-
+
-
| RB31:Sc-indC-Schr_fw || 2013-07-15 || indigoidine exchangeable construct || GAG AAAGAGGAGAAA GGTACC ATGAGCGTAGAGACCATC
+
-
|-
+
-
| RB32:Sc-indC-Schr_rv || 2013-07-15 || indigoidine exchangeable construct || CAT AGATCT GGTTTCCTGTGTGAA TTA TCAGTAGTTGGGCGTCTTG
+
-
|-
+
-
| RB33:entD-MG1655_fw || 2013-07-15 || indigoidine exchangeable construct || CACAGGAAACC GGATCC ATGAAAACTACGCATACCTC
+
-
|-
+
-
| RB34:entD-MG1655_rv || 2013-07-15 || indigoidine exchangeable construct || CAGCGTTATTA GCTAGC TCA TTAATCGTGTTGGCACAGC
+
-
|-
+
-
| RB35:sfp-Naka_fw || 2013-07-15 || indigoidine exchangeable construct || CACAGGAAACC GGATCC ATGAAGATTTACGGAATTTATATGGAC
+
-
|-
+
-
| RB36:sfp-Naka_rv || 2013-07-15 || indigoidine exchangeable construct || CAGCGTTATTA GCTAGC TCA TTATAAAAGCTCTTCGTACGAGAC
+
-
|-
+
-
| RB37:Plum-extr_fw || 2013-07-15 || genomic extraction || ATGTTAGAAAATAATATTACACAATG
+
-
|-
+
-
| RB38:Plum-extr_rv || 2013-07-15 || genomic extraction || TTAGATTATTTTCTCAATCTCAG
+
-
|-
+
-
| RB39:Svert-extr_fw || 2013-07-15 || genomic extraction || GTGATCGCCGCCCTC
+
-
|-
+
-
| RB40:Svert-extr_rv || 2013-07-15 || genomic extraction || TTACGGGACGGCGGTC
+
-
|-
+
-
| RB41:MG1655-extr_fw || 2013-07-15 || genomic extraction || ATGAAAACTACGCATACCTCCCTC
+
-
|-
+
-
| RB42:MG1655-extr_rv || 2013-07-15 || genomic extraction || TTAATCGTGTTGGCACAGCGTTATG
+
-
|-
+
-
| RB43:Bsub-extr_fw || 2013-07-15 || genomic extraction || ATGAAGATTTACGGAATTTATATGGAC
+
-
|-
+
-
| RB44:Bsub-extr_rv || 2013-07-15 || genomic extraction || TTATAAAAGCTCTTCGTACGAGAC
+
-
|-
+
-
| RB45:Sc-indC-Schr_rvG || 2013-07-15 || indigoidine exchangeable construct || CAT GGATCC GGTTTCCTGTGTGAA TTA TCAGTAGTTGGGCGTCTTG
+
-
|- CAGCGTTATTA GCTAGC TCA TTATAAAAGCTCTTCGTACGAGAC
+
-
| RB46:indC-woPPT_rv || 2013-08-05 || indC-pSB1C3 w/o PPTase; RB21 overlap || GCACTATCAGCGTTATTA GCTAGCTCATTA TTA TTAGATTATTTTCTCAATCTCAG
+
-
|-
+
-
| RB47:indC-SpeI_fw || 2013-08-05 || remove cutting sites from indC-Plum || CAAGTTCCTAAACCC ACTAGT CTGGCTTAT ATTATTTATACCTCTGGTAGCAC
+
-
|-
+
-
| RB48:indC-EcoRI_rv || 2013-08-05 || remove cutting sites from indC-Plum || CAATACCCACC GAATTC TTTGAGCT AATTTCTGACAGACAATACC
+
-
|-
+
-
| RB49:indC-EcoRI_fw || 2013-08-05 || remove cutting sites from indC-Plum || AGCTCAAA GAATTC GGTGGGTATTG GGCTTTTTTGTGATC
+
-
|-
+
-
| RB50:indC-SpeI_rv || 2013-08-05 || remove cutting sites from indC-Plum || ATAAGCCAG ACTAGT GGGTTTAGGAACTTG GAACTTGAACTGTG
+
-
|-
+
-
| RB51:pSB(2/3)K3_fw || 2013-08-05 || PPTase plasmid || TAATGA GCTAGC tactagtagcggccgctgcagtc
+
-
|-
+
-
| RB52:pSB(2/3)K3_rv || 2013-08-05 || PPTase plasmid || CAT GGATCC GGTTTCCTGTGTGAA ctctagaagcggccgcgaattcc
+
-
|-
+
-
| RB53:entF-T_fw || 2013-08-07 || bring T-Domain in indC || TGGTGCCATTACATACAGATACT GAAACGATTATCGCCGCGGCATTC
+
-
|-
+
-
| RB54:entF-T_rv || 2013-08-07 || bring T-Domain in indC || AGAGTCTGTCTGTTCAATCCACTT TGCCAGTTTGGCGACAGTTGACG
+
-
|-
+
-
| RB55:tycA1-T_fw || 2013-08-07 || bring T-Domain in indC || TGGTGCCATTACATACAGATACT GAATCGATTCTCGTCTCCATCTGG
+
-
|-
+
-
| RB56:tycA1-T_rv || 2013-08-07 || bring T-Domain in indC || AGAGTCTGTCTGTTCAATCCACTT AGCAACCTGCTCGATCGTCGGGTAATTC
+
-
|-
+
-
| RB57:tycC6-T_fw || 2013-08-07 || bring T-Domain in indC || TGGTGCCATTACATACAGATACT GAACAGCAACTGGCAGCCATCTGGCAAG
+
-
|-
+
-
| RB58:tycC6-T_rv || 2013-08-07 || bring T-Domain in indC || AGAGTCTGTCTGTTCAATCCACTT GGCCACCTTTTCGATCGTCGGATACTG
+
-
|-
+
-
| RB59:delH4-T_fw || 2013-08-07 || bring T-Domain in indC || TGGTGCCATTACATACAGATACT GAGACGCTGCTGGCCCGTATCTG
+
-
|-
+
-
| RB60:delH4-T_rv || 2013-08-07 || bring T-Domain in indC || AGAGTCTGTCTGTTCAATCCACTT CGCCAGCTCGGCAATGCTTTGC
+
-
|-
+
-
| RB61:delH5-T_fw || 2013-08-07 || bring T-Domain in indC || TGGTGCCATTACATACAGATACT GCCATGGCGCTGGCCCGCATC
+
-
|-
+
-
| RB62:delH5-T_rv || 2013-08-07 || bring T-Domain in indC || AGAGTCTGTCTGTTCAATCCACTT CAGCAGGCCGGCAATGGTCG
+
-
|-
+
-
| RB63:pSB(2/3)K3_rv_korr || 2013-08-07 || PPTase plasmid || CAT GGATCC GGTTTCCTGTGTGAA CTCTAGTATGTGTGAAATTGTTATCC
+
-
|-
+
-
| RB64:DelC-extr_fw || 2013-08-07 || get Delftia PPTase || ATGCAGCTCGTGTCCGTGCGTGAG
+
-
|-
+
-
| RB65:DelC-extr_rv || 2013-08-07 || get Delftia PPTase || TCATGTCGATTCCTTGGTGCGCCAC
+
-
|-
+
-
| RB66:DelC-pSB2K3_fw || 2013-08-07 || DelC in PPTase-construct || CACAGGAAACC GGATCC ATGCAGCTCGTGTCCGTGCGTGAG
+
-
|-
+
-
| RB67:DelC-pSB2K3_rv || 2013-08-07 || DelC in PPTase-construct || CAGCGTTATTA GCTAGC TCA TCATGTCGATTCCTTGGTGCGCCAC
+
-
|-
+
-
| RB68:indC-SpeI_fw_corr || 2013-08-09 || indC w/o cutting sites corrected || CAAGTTCCTAAACCC ACgAGT CTGGCTTAT ATTATTTATACCTCTGGTAGCAC
+
-
|-
+
-
| RB69:indC-EcoRI_rv_corr || 2013-08-09 || indC w/o cutting sites corrected || CAATACCCACC GAAcTC TTTGAGCT AATTTCTGACAGACAATACC
+
-
|-
+
-
| RB70:indC-EcoRI_fw_corr || 2013-08-09 || indC w/o cutting sites corrected || AGCTCAAA GAgTTC GGTGGGTATTG GGCTTTTTTGTGATC
+
-
|-
+
-
| RB71:indC-SpeI_rv_corr || 2013-08-09 || indC w/o cutting sites corrected || ATAAGCCAG ACTcGT GGGTTTAGGAACTTG GAACTTGAACTGTG
+
-
|-
+
-
| RB72:bpsA-TTE_rv || 2013-08-09 || exchange T and TE-domain || GCACTATCAGCGTTATTA GCTAGC TCATTA TTCCCCCAGCAGGTATCTAATATG
+
-
|-
+
-
| RB73:entF-TTE_rv || 2013-08-09 || exchange T and TE-domain || GCACTATCAGCGTTATTA GCTAGC TCA TTACCTGTTTAGCGTTGCGCGAATAATCG
+
-
|-
+
-
| RB74:tycC6-TTE_rv || 2013-08-09 || exchange T and TE-domain || GCACTATCAGCGTTATTA GCTAGC TCA TTATTTCAGGATGAACAGTTCTTGC
+
-
|-
+
-
| RB75:delH5-TTE_rv || 2013-08-09 || exchange T and TE-domain || GCACTATCAGCGTTATTA GCTAGC TCATTA TCAGTCCAGCGCGTACTCCAG
+
-
|-
+
-
| RB76:plu2670-T_fw || 2013-08-15 || bring T-Domain in indC || TGGTGCCATTACATACAGATACT GAAACCACACTGGCTGCTATC
+
-
|-
+
-
| RB77:plu2670-T_rv || 2013-08-15 || bring T-Domain in indC || AGAGTCTGTCTGTTCAATCCACTT CGCAAATGCGGATAACACC
+
-
|-
+
-
| RB78:indC-SpeI_rv_corr2 || 2013-08-26 || indC w/o cutting sites w/o frameshift || ATAAGCCAG ACTcGT GGGTTTAGGAACTTG AACTGTG
+
-
|-
+
-
| RB79:indC_woPPTase_rv2 || 2013-08-29 || indC-pSB1C3 w/o PPTase; RB35 5' part || CAGCGTTATTA GCTAGC TCA TTAGATTATTTTCTCAATCTCAG
+
-
|-
+
-
| RB80:indCdT_3_fw || 2013-09-06 || || GGCGGATATCCTATGAGTTTGAGATTGC
+
-
|-
+
-
| RB81:indCdT_3_rv || 2013-09-06 || || TCCATTGGCCGTCAAAGGTAATTTATCG
+
-
|-
+
-
| RB82:bpsA-T3_fw || 2013-09-06 || || CGATAAATTACCTTTGACGGCCAATGGA AAGATCGATGTGAAAGCACTGGCCGCTTCTGACC
+
-
|-
+
-
| RB83:bpsA-T3_rv || 2013-09-06 || || GCAATCTCAAACTCATAGGATATCCGCC CAGACCTGGCCAGCAGATCACAGG
+
-
|-
+
-
| RB84:indCdT_4_fw || 2013-09-06 || || TTTGAAGTTGCATACCAGCTTGAACAAGC
+
-
|-
+
-
| RB85:bpsA-T4_rv || 2013-09-06 || || GCTTGTTCAAGCTGGTATGCAACTTCAAA TGCCACGCGAGCTCCGAAGCTATATCC
+
-
|-
+
-
| RB86:indC-T2_fw || 2013-09-06 || || TTACAGATACTTTTTCAATCTCCTAATATAGCAG
+
-
|-
+
-
| RB87:indC-T2_rv || 2013-09-06 || || TACTTCCATCCAAATTTTTCCAAGC
+
-
|-
+
-
| RB88:bpsA-T2_fw || 2013-09-06 || || GCTTGGAAAAATTTGGATGGAAGTA CTGAGACGCGAAAATGCTAGTGTCC
+
-
|-
+
-
| RB89:bpsA-T2_rv || 2013-09-06 || || TTAGGAGATTGAAAAAGTATCTGTAA AGGCAGGGACACTCCCAGTCTAGCATTCAG
+
-
|-
+
-
| RB90:indC-T-screen_fw || 2013-09-13 || || GATCAATGCGGCCTTTAATATTCG
+
-
|-
+
-
| RB91:bpsA-T-screen_fw || 2013-09-13 || || GGAACTGAATGCTAGACTGGGAG
+
-
|-
+
-
| RB92:entF-T3_fw || 2013-09-16 || || CGATAAATTACCTTTGACGGCCAATGGA AAGCTGGATCGCAAAGCCTTACCGTTG
+
-
|-
+
-
| RB93:tycA1-T3_fw || 2013-09-16 || || CGATAAATTACCTTTGACGGCCAATGGA AAGATCGACCGCAAAGCGTTGC
+
-
|-
+
-
| RB94:tycC6-T3_fw || 2013-09-16 || || CGATAAATTACCTTTGACGGCCAATGGA AAAGTGGATCGCAAAGCTTTG
+
-
|-
+
-
| RB95:delH4-T3_fw || 2013-09-16 || || CGATAAATTACCTTTGACGGCCAATGGA AAGCTGGACCGGCAGGCCCTG
+
-
|-
+
-
| RB96:delH5-T3_fw || 2013-09-16 || || CGATAAATTACCTTTGACGGCCAATGGA AAGCTTGACCGTGGTGCGCTG
+
-
|-
+
-
| RB97:plu2642-T3_fw || 2013-09-16 || || CGATAAATTACCTTTGACGGCCAATGGA AAAATCGATTTCGACACATTACAAG
+
-
|-
+
-
| RB98:plu2670-T3_fw || 2013-09-16 || || CGATAAATTACCTTTGACGGCCAATGGA AAGCTGGACCGTCGGGCGTTACCGGCAC
+
-
|-
+
-
| RB99:plu2642_pRB23-plu_fw || 2013-09-24 || plu2642 on pSB1C3 || CATACTAGAG AAAGAGGAGAAA GGTACC ATGCAATCAACTCTCCCAATAATAAAATGG
+
-
|-
+
-
| RB100:plu2642_pRB23-plu_rv || 2013-09-24 || plu2642 on pSB1C3 || CAGCGTTATTA GCTAGC TCA TCAACTCAAGAATCGAGCTAATTCGTTAACAC
+
-
|-
+
-
| RB101:plu2642_A_rv || 2013-09-24 || || GTTCTGGCATCCTTCTTAATTAACAT TGCAGCGTGTTTCACTTGATCGATAAC
+
-
|-
+
-
| RB102:indC_Ox_fw || 2013-09-24 || || ATGTTAATTAAGAAGGATGCCAGAAC
+
-
|-
+
-
| RB103:plu2642_valInd_fw || 2013-09-24 || || CAGCCTGCCAGCAGCAAAACG ATGCAATCAACTCTCCCAATAATAAAATGG
+
-
|-
+
-
| RB104:plu2642_valInd_rv || 2013-09-24 || || CAT CGTTTTGCTGCTGGCAGGCTG
+
-
|-
+
-
| RB105:tycC2_T3_fw || 2013-09-24 || || CGATAAATTACCTTTGACGGCCAATGGA AAAGTGGATCGCAAGGCATTG
+
-
|-
+
-
| RB106:tycC2_T1_rv || 2013-09-24 || || AGAGTCTGTCTGTTCAATCCACTT GGCCAGGCCCGCGATCGTTGG
+
-
|-
+
-
| RB107:tycC2_A_fw || 2013-09-24 || || CATACTAGAG AAAGAGGAGAAA GGTACC ATGACGGAAGCGGAAAAACGCACACTCCTTC
+
-
|-
+
-
| RB108:tycC2_A_rv || 2013-09-24 || || GTTCTGGCATCCTTCTTAATTAACAT GACGACCGCTTTGACCGCTTCATG
+
-
|-
+
-
| RB109:tycC2_valInd_fw || 2013-09-24 || || CAGCCTGCCAGCAGCAAAACG ATGACGGAAGCGGAAAAACGCACACTCCTTC
+
-
|-
+
-
| RB110:plu2642_TE_fw || || || AGTGGATTGAACAGACAGACTCT AAAATCGTTGAAGGTGAAGTAACCAG
+
-
|-
+
-
|}
+
-
<br/>
+
-
=Plasmids=
 
-
==Delftibactin==
 
{|class="wikitable sortable"
{|class="wikitable sortable"
|-
|-
!Name!!Date!!Brief Description!!Genotype!!Plasmid Map!!GenBank-File
!Name!!Date!!Brief Description!!Genotype!!Plasmid Map!!GenBank-File
|-
|-
-
|pIK1||2013-08-12||Methylmalonyl-CoA pathway with permeability device and sfp||pSB3C5-lacP-BBa_B0034-BBa_I746200-BBa_B0029-pcc-acca2-BBa_B0030-sfp||[[File:Heidelberg_PIK1.png|160px|pIK1]]||[[media:Heidelberg_PIK1.gb.txt|pIK1]]
 
-
|-
 
-
|pIK2||2013-08-12||Methylmalonyl-CoA pathway with sfp||pSB3C5-lacP-BBa_B0034-pcc-acca2-BBa_B0030-sfp||[[File:Heidelberg_PIK2.png|160px|pIK2]]||[[media:Heidelberg_PIK2.gb.txt|pIK2]]
 
-
|-
 
-
|pIK7||2013-08-28||Methylmalonyl-CoA pathway with sfp||pSB3K3-lacP-BBa_B0034-pcc-acca2-BBa_B0030-sfp||[[File:Heidelberg_PIK7.png|160px|pIK7]]||[[media:Heidelberg_PIK7.gb.txt|pIK7]]
 
-
|-
 
-
|pIK8||2013-09-12||Methylmalonyl-CoA pathway with sfp and the permeability device behind a weak promoter||pSB3C5-lacP-BBa_B0034-pcc-acca2-BBa_B0030-sfp-BBa_B0010-BBa_B0030-BBa_I746200-BBa_B0012||[[File:Heidelberg_PIK8.png|160px|pIK8]]||[[media:Heidelberg_PIK8.gb.txt|pIK8]]
 
-
|-
 
-
|pIK9||2013-09-18||Methylmalonyl-CoA pathway with sfp and the permeability device behind a weak promoter||pSB3K3-lacP-BBa_B0034-pcc-acca2-BBa_B0030-sfp-BBa_B0010-BBa_B0030-BBa_I746200-BBa_B0012||[[File:Heidelberg_PIK9.png|160px|pIK9]]||[[media:Heidelberg_PIK9.gb.txt|pIK9]]
 
-
|-
 
-
|pIK10||2013-09-18||Methylmalonyl-CoA pathway with sfp and the permeability device behind a weak promoter, for submission||pSB1C3-lacP-BBa_B0034-pcc-acca2-BBa_B0030-sfp-BBa_B0010-BBa_B0030-BBa_I746200-BBa_B0012||[[File:Heidelberg_PIK10.png|160px|pIK10]]||[[media:Heidelberg_PIK10.gb.txt|pIK10]]
 
-
|-
 
-
|pIK11||2013-09-18||Methylmalonyl-CoA pathway with sfp, for submission||pSB1C3-lacP-BBa_B0034-pcc-acca2-BBa_B0030-sfp||[[File:Heidelberg_PIK11.png|160px|pIK11]]||[[media:Heidelberg_PIK11.gb.txt|pIK11]]
 
-
|-
 
-
|pHM01||2013-04-29|| DelH construct assembled by Restriction Ligation Strategy ||pSB6A1-BBa_K206000(AraC)-DelH-BBa_I732019(lacZ)-B0015(stop-codon) ||[[File:Heidelberg_Res-DelH-finalplasmid.png|160px|pHM01]]||[[media:Heidelberg_DelH-pSB6A1-AraC-lacZ.gb.txt|pHM01]]
 
-
|-
 
-
|pHM02||2013-07-08|| DelH construct assembled by Gibson Assemby ||pSB6A1-R0010(lacI)-B0035(RBS)-DelH-B0034(RBS)-E1010(mRFP)-B0015(stop) ||[[File:Heidelberg_FinalPlasmid.png|160px|pHM02]]||[[media:Heidelberg_DelH-BB(Gibson-pSB6A1-AraC-lacZ).gb.txt|pHM02]]
 
-
|-
 
-
|pHM03||2013-04-29|| DelH construct assembled by Gibson Assemby for existing backbone ||pSB6A1-R0010(lacI)-B0034(RBS)-DelH-B0035(RBS)-E1010(mRFP)-B0015(stop) ||[[File:Heidelberg_Construct-plasmid.png|160px|pHM03]]||[[media:Heidelberg_PSB6A1+DelH.gb.txt|pHM03]]
 
-
|-
 
-
|pHM04||2013-08-12|| DelH construct assembled by Gibson Assemby without mRFP ||pSB6A1-R0010(lacI)-B0034(RBS)-DelH-B0015(stop) ||[[File:Heidelberg_PHM 04.png|160px|pHM04]]||[[media:Heidelberg_PHM04-DelH-pSB6A1(without mRFP).gb.txt|pHM04]]
 
-
|-
 
-
|pHM05||2013-08-12|| DelH construct assembled by Gibson Assemby with tetracycline resistance, without mRFP|| pSB6A1-R0010(lacI)-DelH-B0015(stop)-pSB1T3(tetR) ||[[File:Heidelberg_PHM05-DelH-tetR-pSB6A1.png|160px|pHM05]]||[[media:Heidelberg_PHM05-DelH-tetR-pSB6A1.gb.txt|pHM05]]
 
-
|-
 
-
|pFSN||2013-06-28||DelRest, Delftibactin cluster from ''D. Acidovorans'' ||pSB4K5-lacZ-BBa_B0035-DelA-DelB-DelC-DelE-DelF-DelG-DelO-DelP-DelL-BBa_B0034||[[File:Heidelberg_PFSN.png|160px|pFSN]]||[[media:Heidelberg_Psb4k5+BBa J04450 DelRest.gb.txt|pFSN]]
 
-
|-
 
-
|pFS_02||2013-09-26||DelH construct assembled by Gibson Assemby with weak Promotor, weak RBS ||pSB6A1-BBa_J23114-BBa_B0032-DelH||[[File:Heidelberg_pFS_02.png|160px|pFS_02]]||[[media:Heidelberg_PSB6A1 BBa J23114 BBa B0032 DelH.gb.txt|pFS_02]]
 
-
|-
 
-
|pFS_03||2013-09-26|| Helper plasmid for DelH restriction cloning, DelH, flanked by KpnI and BamHI site, without promotor,||pSB4K5-DelH||[[File:Heidelberg_pFS_03.png|160px|pFS_03]]||[[media:Heidelberg_PSB4K5min KpnI DelH BamHI.gb.txt|pFS_03]]
 
-
|-
 
-
|pFS_04||2013-09-26||Target plasmid for DelH restriction cloning, ccdB cassette, flanked by KpnI and BamHI site ||pSB6A1-BBa_J23114-BBa_B0032-ccdBcassette||[[File:Heidelberg_pFS_02.png|160px|pFS_02]]||[[media:Heidelberg_PSB6A1 BBa J23114 BBa B0032 ccdB cassette.gb.txt|pFS_04]]
 
-
|-
 
-
|pFS_05||2013-09-26|| DelH construct assembled by restriction cloning using pFS_03 and pFS-04 DelH, flanked by KpnI and BamHI site, under weak promotor,||pSB6A1-BBa_J23114-BBa_B0032-DelH||[[File:Heidelberg_pFS_03.png|160px|pFS_05]]||[[media:Heidelberg_DELH final pSB6A1 BBa J23114 BBa B0032 KpnI DelH BamHI.gb.txt|pFS_05]]
 
|}
|}
-
<br/>
+
 
-
==Module Shuffling==
+
-
{|class="wikitable sortable"
+
-
|-
+
-
!Name!!Date!!Brief Description!!Genotype!!Plasmid Map!!GenBank-File
+
-
|-
+
-
|pIK03||2013-mm-dd||NRPS for Pro-Leu||pSB1C3+TycB1dComdC+TycC6||[[File:Heidelberg_Dipeptide.png|160px|pIK03]]||[[media:Heidelberg_PSB1C3+TycB1dComdC+TycC6.gb.txt|pIK03]]
+
-
|-
+
-
|pIK04||2013-mm-dd||NRPS for Phe-Orn-Leu||pSB1C3+TycAdCom+TycC5-TycC6||[[File:Heidelberg_Tripeptide_I.png|160px|pIK04]]||[[media:Heidelberg_PSB1C3+TycAdCom+TycC5-TycC6.gb.txt|pIK04]]
+
-
|-
+
-
|pIK05||2013-mm-dd||NRPS for Pro-Orn-Leu||pSB1C3+TycB1dComdC+TycC5-TycC6||[[File:Heidelberg_Tripeptide_II.png|160px|pIK05]]||[[media:Heidelberg_PSB1C3+TycBdComdC+TycC5-TycC6.gb.txt|pIK05]]
+
-
|-
+
-
|pPW01||2013-mm-dd||NRPS for Orn-Pro-Phe-Leu||pSB1C3+TycC5+TycB1dCom-C(TycB2)+TycAdCom+TycC6||[[File:Heidelberg_Tetrapeptide_I.png|160px|pPW01]]||[[media:Heidelberg_Bla.gb.txt|pPW01]]
+
-
|-
+
-
|pPW02||2013-mm-dd||NRPS for Orn-Pro-Phe-Leu||pSB1C3+TycC5+TycB1dCom-C(TycB2)+TycAdE+TycC6||[[File:Heidelberg_Tetrapeptide_II.png|160px|pPW02]]||[[media:Heidelberg_Blaa.gb.txt|pPW02]]
+
-
|-
+
-
|pPW03||2013-08-18||NRPS for Phe-Indigoidine||pSB1C3+TycA+C(TycC2)+IndC||[[File:Heidelberg_Construct 1.png|160px|pPW03]]||[[media:Heidelberg_PSB1C3+TycAdE+C(TycC2)+indC.gb.txt|pPW03]]
+
-
|-
+
-
|pPW04||2013-08-18||NRPS for Asn-Indigoidine||pSB1C3+TycC1-C(TycC2)+IndC||[[File:Heidelberg_Construct 2.png|160px|pPW04]]||[[media:Heidelberg_PSB1C3+TycC1-C(C2)+indC.gb.txt|pPW04]]
+
-
|-
+
-
|pPW05||2013-08-18||NRPS for Val-Indigoidine||pSB1C3+TycC4+C(TycC2)+IndC||[[File:Heidelberg_Construct 3.png|160px|pPW05]]||[[media:Heidelberg_PSB1C3+TycC4+C(TycC2)+indC.gb.txt|pPW05]]
+
-
|-
+
-
|pPW06||2013-09-07||NRPS for Orn-Val-Indigoidine||pSB1C3+TycC5dC+C(TycC4)+TycC4dC+C(TycC2)+indC||[[File:Heidelberg_Construct A.png|160px|pPW06]]||[[media:Heidelberg_A_pSB1C3+TycC5dC+C(TycC4)+TycC4dC+C(TycC2)+indC.gb.txt|pPW06]]
+
-
|-
+
-
|pPW07||2013-09-07||NRPS for Orn-Val-Val-Indigoidine||pSB1C3+TycC5dC+TycC4+C(TycC4)+TycC4dC+C(TycC2)+indC||[[File:Heidelberg_Construct B.png|160px|pPW07]]||[[media:Heidelberg_B_pSB1C3+TycC5dC+TycC4+C(TycC4)+TycC4dC+C(TycC2)+indC.gb.txt|pPW07]]
+
-
|-
+
-
|pPW08||2013-09-07||NRPS for Val-Val-Indigoidine||pSB1C3+TycC4dC+C(TycC4)+TycC4dC+C(TycC2)+indC||[[File:Heidelberg_Construct C.png|160px|pPW08]]||[[media:Heidelberg_C_pSB1C3+TycC4dC+C(TycC4)+TycC4dC+C(TycC2)+indC.gb.txt|pPW08]]
+
-
|-
+
-
|pPW09||2013-09-07||NRPS for Pro-Leu-Indigoidine||pSB1C3+TycB1dComdC+TycC6dTE+C(TycC2)+indC||[[File:Heidelberg_Construct D.png|160px|pPW09]]||[[media:Heidelberg_D_pSB1C3+TycB1dComdC+TycC6dTE+C(TycC2)+indC.gb.txt|pPW09]]
+
-
|-
+
-
|pPW10||2013-09-07||NRPS for Pro-Leu-Val-Indigoidine||pSB1C3+TycB1dComdC+TycC6dTE+C(TycC4)+TycC4dC+C(TycC2)+indC||[[File:Heidelberg_Construct E.png|160px|pPW09]]||[[media:Heidelberg_E_pSB1C3+TycB1dComdC+TycC6dTE+C(TycC4)+TycC4dC+C(TycC2)+indC.gb.txt|pPW10]]
+
-
|-
+
-
|pPW11||2013-09-07||NRPS for Phe-Orn-Leu-Indigoidine||pSB1C3+TycAdCom+TycC5-TycC6dTE+C(TycC2)+indC||[[File:Heidelberg_Construct F.png|160px|pPW11]]||[[media:Heidelberg_F_pSB1C3+TycAdCom+TycC5-TycC6dTE+C(TycC2)+indC.gb.txt|pPW11]]
+
-
|-
+
-
|pPW12||2013-09-07||NRPS for Phe-Orn-Leu-Val-Indigoidine||pSB1C3+TycAdCom+TycC5-TycC6dTE+C(TycC4)+TycC4dC+C(TycC2)+indC||[[File:Heidelberg_Construct G.png|160px|pPW12]]||[[media:Heidelberg_G_pSB1C3+TycAdCom+TycC5-TycC6dTE+C(TycC4)+TycC4dC+C(TycC2)+indC.gb.txt|pPW12]]
+
-
|-
+
-
|pJS01||2013-09-18||Helper plasmid for the tagging of NRPs with Indigoidine||pSB1C3+ccdB+C(TycC2)+indC||[[File:Heidelberg_Construct ccdB.png|160px|pJS01]]||[[media:Heidelberg_PSB1C3+ccdB+c(TycC2)+indC.gb.txt|pJS01]]
+
-
|}
+
-
<br/>
+
-
==Linker variation==
+
-
{|class="wikitable sortable"
+
-
|-
+
-
!Name!!Date!!Brief Description!!Genotype!!Plasmid Map!!GenBank-File
+
-
|-
+
-
|pAR01||2013-16-09|| NRPS for Valin-Asparagine-Indigoidine-Expression with double short linkers || pSB1C3-TycC4dCom_s+TycC2_s+indC || [[File:Heidelberg_ ConstructLV1.png|160px|pAR01]]|| [[media:Heidelberg_PSB1C3+TycC4_s+C(TycC2)_s+indC.gb|pAR01]]
+
-
|-
+
-
|pAR02||2013-16-09||  NRPS for Valin-Asparagine-Indigoidine-Expression with C4 short linker || pSB1C3-TycC4dCom_s+TycC2_m+indC || [[File:Heidelberg_ ConstructLV2.png|160px|pAR03]]|| [[media:Heidelberg_PSB1C3+TycC4_s+C(TycC2)_m+indC.gb|pAR02]]
+
-
|-
+
-
|pAR03||2013-16-09||  NRPS for Valin-Asparagine-Indigoidine-Expression with C2 short linker || pSB1C3-TycC4dCom_m+TycC2_s+indC || [[File:Heidelberg_ ConstructLV3.png|160px|pAR03]]|| [[media:Heidelberg_PSB1C3+TycC4_m+C(TycC2)_s+indC.gb|pAR03]]
+
-
|-
+
-
|pAR04||2013-16-09||  NRPS for Valin-Asparagine-Indigoidine-Expression with medium linkers || pSB1C3-TycC4dCom+TycC2+indC || [[File:Heidelberg_ ConstructLV4.png|160px|pAR04]]|| [[media:Heidelberg_PSB1C3+TycC4_m+C(TycC2)_m+indC.gb|pAR04]]
+
-
|-
+
-
|pAR05||2013-16-09||  NRPS for Valin-Asparagine-Indigoidine-Expression with C2 long linker || pSB1C3-TycC4dCom_m+TycC2_l+indC || [[File:Heidelberg_ ConstructLV5.png|160px|pAR05]]|| [[media:Heidelberg_PSB1C3+TycC4_m+C(TycC2)_l+indC.gb|pAR05]]
+
-
|-
+
-
|pAR06||2013-16-09||  NRPS for Valin-Asparagine-Indigoidine-Expression with C4 long linker || pSB1C3-TycC4dCom_l+TycC2_m+indC || [[File:Heidelberg_ ConstructLV6.png|160px|pAR06]]|| [[media:Heidelberg_PSB1C3+TycC4_l+C(TycC2)_m+indC.gb|pAR06]]
+
-
|-
+
-
|pAR07||2013-16-09||  NRPS for Valin-Asparagine-Indigoidine-Expression with long linkers || pSB1C3-TycC4dCom_l+TycC2_l+indC || [[File:Heidelberg_ ConstructLV7.png|160px|pAR07]]|| [[media:Heidelberg_PSB1C3+TycC4_l+C(TycC2)_l+indC.gb|pAR07]]
+
-
|-
+
-
|pAR08||2013-16-09||  NRPS for Valin-Asparagine-Indigoidine-Expression with C4 short and C2 long linker || pSB1C3-TycC4dCom_s+TycC2_l+indC || [[File:Heidelberg_ ConstructLV8.png|160px|pAR08]]|| [[media:Heidelberg_PSB1C3+TycC4_s+C(TycC2)_l+indC.gb|pAR08]]
+
-
|-
+
-
|pAR09||2013-16-09||  NRPS for Valin-Asparagine-Indigoidine-Expression with C4 long and C2 short linker || pSB1C3-TycC4dCom_l+TycC2_s+indC || [[File:Heidelberg_ ConstructLV9.png|160px|pAR09]]|| [[media:Heidelberg_PSB1C3+TycC4_l+C(TycC2)_s+indC.gb|pAR09]]
+
-
|-
+
-
|}
+
-
<br/>
+
-
==Dinner plasmids==
+
-
{|class="wikitable sortable"
+
-
|-
+
-
!Name!!Date!!Brief Description!!Genotype!!Plasmid Map!!GenBank-File
+
-
|-
+
-
|<div id="pIK6">pIK6</div>||2013-08-27||pIK6: special Ilia plasmid. This plasmid does not fullfill iGEM's RFC10 but is nevertheless special. It is for expression of tasty Ham driven by a strong lacCheese inducible promoter, a RBSalami for increased fleshiness, which can only terminated by a Champignon stop codon. The pIZa backbone with Pepper resistance and high copy Onion ori will improve the users taste stimulation. Bon Appétit! ||pIZa-lacCheese P-RBSalami-ham CDS-Champ Term-Pepper res-Onion Ori||[[File:Heidelberg_PIK6.png|160px|pIK6]]||[[media:Heidelberg_PIK6.txt|pIK6]]
+
-
|-
+
-
|}
+
-
<br/>
+
-
==Domain Shuffling and PPTases==
+
-
{|class="wikitable sortable"
+
-
|-
+
-
!Name!!Date!!Brief Description!!Genotype!!Plasmid Map!!GenBank-File
+
-
|-
+
-
|<div id="pMM64">pMM64</div>||2013-06-01||bpsA(fus)||pETDuet-1-T7lac-bpsA(fus)||[[File:Heidelberg_PMM064.png|160px|pMM64]]||[[media:Heidelberg_PMM064.gb.txt|pMM64]]
+
-
|-
+
-
|<div id="pMM65">pMM65</div>||2013-06-01||svp(fus)||pETDuet-1-T7lac-svp(fus)||[[File:Heidelberg_PMM065.png|160px|pMM65]]||[[media:Heidelberg_PMM065.gb.txt|pMM65]]
+
-
|-
+
-
|<div id="pKH4">pKH4</div>||2013-09-01||pRB3/pMM64 derived plasmid without sfp||pSB1C3-lacP-BBa_B0034-KpnI-indC-BBa_B0029-nonsense||[[File:Heidelberg_PKH4.png|160px|pKH4]]||[[media:Heidelberg_PKH4.gb|pKH4]]
+
-
|-
+
-
|<div id="pKH6">pKH6</div>||2013-09-05||BBa CDS sfp only for part submission||pSB1C3-sfp||[[File:Heidelberg_PKH6.png|160px|pKH6]]||[[media:Heidelberg_PKH6.gb|pKH6]]
+
-
|-
+
-
|<div id="pKH7">pKH7</div>||2013-09-05||BBa CDS svp only for part submission||pSB1C3-svp||[[File:Heidelberg_PKH7.png|160px|pKH7]]||[[media:Heidelberg_PKH7.gb|pKH7]]
+
-
|-
+
-
|<div id="pKH8">pKH8</div>||2013-09-05||BBa CDS entD only for part submission||pSB1C3-entD||[[File:Heidelberg_PKH8.png|160px|pKH8]]||[[media:Heidelberg_PKH8.gb|pKH8]]
+
-
|-
+
-
|<div id="pKH9">pKH9</div>||2013-09-05||BBa CDS delC only for part submission||pSB1C3-delC||[[File:Heidelberg_PKH9.png|160px|pKH9]]||[[media:Heidelberg_PKH9.gb|pKH9]]
+
-
|-
+
-
|<div id="pRB1">pRB1</div>||2013-07-01||bpsA svp(pMM65)||pSB1C3-lacP-BBa_B0034-bpsA-BBa_B0029-svp(pMM65)||N.A||N.A
+
-
|-
+
-
|<div id="pRB2">pRB2</div>||2013-07-08||bpsA svp||HindIII-pSB1C3-lacP-BBa_B0034-bpsA-BBa_B0029-HindIII-svp||N.A||N.A
+
-
|-
+
-
|<div id="pRB3">pRB3</div>||2013-07-15||indC sfp||NheI-pSB1C3-lacP-BBa_B0034-KpnI-indC-BBa_B0029-BamHI-sfp||[[File:Heidelberg_pRB03.png|160px|pRB03]]||[[media:Heidelberg_PRB03.gb|pRB03]]
+
-
|-
+
-
|<div id="pRB4">pRB4</div>||2013-07-15||indC svp||NheI-pSB1C3-lacP-BBa_B0034-KpnI-indC-BBa_B0029-BamHI-svp||[[File:Heidelberg_pRB04.png|160px|pRB04]]||[[media:Heidelberg_PRB04.gb|pRB04]]
+
-
|-
+
-
|<div id="pRB5">pRB5</div>||2013-07-15||indC svpF||NheI-pSB1C3-lacP-BBa_B0034-KpnI-indC-BBa_B0029-BamHI-svp(pMM65)||[[File:Heidelberg_pRB05.png|160px|pRB05]]||[[media:Heidelberg_PRB05.gb|pRB05]]
+
-
|-
+
-
|<div id="pRB6">pRB6</div>||2013-07-15||indC entD||NheI-pSB1C3-lacP-BBa_B0034-KpnI-indC-BBa_B0029-BamHI-entD||[[File:Heidelberg_pRB06.png|160px|pRB06]]||[[media:Heidelberg_PRB06.gb|pRB06]]
+
-
|-
+
-
|<div id="pRB7">pRB7</div>||2013-07-15||bpsA(pMM64) sfp||NheI-pSB1C3-lacP-BBa_B0034-KpnI-bpsA(pMM64)-BBa_B0029-BamHI-sfp||[[File:Heidelberg_pRB07.png|160px|pRB07]]||[[media:Heidelberg_PRB07.gb|pRB07]]
+
-
|-
+
-
|<div id="pRB8">pRB8</div>||2013-07-15||bpsA(pMM64) svpF||NheI-pSB1C3-lacP-BBa_B0034-KpnI-bpsA(pMM64)-BBa_B0029-BamHI-svp(pMM65)||[[File:Heidelberg_pRB08.png|160px|pRB08]]||[[media:Heidelberg_PRB08.gb|pRB08]]
+
-
|-
+
-
|<div id="pRB9">pRB9</div>||2013-07-15||bpsA(pMM64) svp||NheI-pSB1C3-lacP-BBa_B0034-KpnI-bpsA(pMM64)-BBa_B0029-BamHI-svp||[[File:Heidelberg_pRB09.png|160px|pRB09]]||[[media:Heidelberg_PRB09.gb|pRB09]]
+
-
|-
+
-
|<div id="pRB10">pRB10</div>||2013-07-15||bpsA(pMM64) entD||NheI-pSB1C3-lacP-BBa_B0034-KpnI-bpsA(pMM64)-BBa_B0029-BamHI-entD||[[File:Heidelberg_pRB10.png|160px|pRB10]]||[[media:Heidelberg_PRB10.gb|pRB10]]
+
-
|-
+
-
|<div id="pRB11">pRB11</div>||2013-07-29||pKH1-der bpsA(pMM64)-ccdb svpF||pSB1C3-lacP-BBa_B0034-bpsA(pMM64)(ccdb)-BBa_B0029-svp(pMM65)||N.A||N.A
+
-
|-
+
-
|<div id="pRB12">pRB12</div>||2013-07-29||pKH2-der bpsA(pMM64)-ccdb svpF||pSB1C3-lacP-BBa_B0034-bpsA(pMM64)(ccdb)-BBa_B0029-svp(pMM65)||N.A||N.A
+
-
|-
+
-
|<div id="pRB13">pRB13</div>||2013-07-29||pRB3-der indC-ccdb sfp||NheI-pSB1C3-lacP-BBa_B0034-KpnI-indC(ccdb)-BBa_B0029-BamHI-sfp||N.A||N.A
+
-
|-
+
-
|<div id="pRB14">pRB14</div>||2013-08-12||pRB3-der indC-ccdB sfp||NheI-pSB1C3-lacP-BBa_B0034-KpnI-indC(ccdB)-BBa_B0029-BamHI-sfp||N.A||N.A
+
-
|-
+
-
|<div id="pRB15">pRB15</div>||2013-08-12||pSB3K3 sfp||pSB3K3-lacP-BBa_B0029-BamHI-sfp-NheI||[[File:Heidelberg_PRB15.png|160px|pRB15]]||[[media:Heidelberg_PRB15.gb|pRB15]]
+
-
|-
+
-
|<div id="pRB16">pRB16</div>||2013-08-12||pSB3K3 svp||pSB3K3-lacP-BBa_B0029-BamHI-svp-NheI||[[File:Heidelberg_PRB16.png|160px|pRB16]]||[[media:Heidelberg_PRB16.gb|pRB16]]
+
-
|-
+
-
|<div id="pRB17">pRB17</div>||2013-08-12||pSB3K3 entD||pSB3K3-lacP-BBa_B0029-BamHI-entD-NheI||[[File:Heidelberg_PRB17.png|160px|pRB17]]||[[media:Heidelberg_PRB17.gb|pRB17]]
+
-
|-
+
-
|<div id="pRB18">pRB18</div>||2013-08-12||pSB3K3 delC||pSB3K3-lacP-BBa_B0029-BamHI-delC-NheI||[[File:Heidelberg_pRB18.png|160px|pRB18]]||[[media:Heidelberg_pRB18.gb|pRB18]]
+
-
|-
+
-
|<div id="pRB19">pRB19</div>||2013-08-19||pRB14-der indC-ccdB||NheI-pSB1C3-lacP-BBa_B0034-KpnI-indC(ccdB)||N.A||N.A
+
-
|-
+
-
|<div id="pRB20">pRB20</div>||2013-08-19||pRB19-der indC-HD-ccdB||NheI-pSB1C3-lacP-BBa_B0034-KpnI-indC-HD(ccdB)||N.A||N.A
+
-
|-
+
-
|<div id="pRB21">pRB21</div>||2013-08-19||pSB1C3 indC||NheI-pSB1C3-lacP-BBa_B0034-KpnI-indC||[[File:Heidelberg_PRB21.png|160px|pRB21]]||[[media:Heidelberg_PRB22.gb|pRB21]]
+
-
|-
+
-
|<div id="pRB22">pRB22</div>||2013-08-19||pRB21-der indC-HD||NheI-pSB1C3-lacP-BBa_B0034-KpnI-indC-HD||[[File:Heidelberg_PRB22.png|160px|pRB22]]||[[media:Heidelberg_PRB22.gb|pRB22]]
+
-
|-
+
-
|<div id="pRB23">pRB23</div>||2013-08-19||pRB22-der indC-HD||NheI-pSB1C3-lacP-BBa_B0034-KpnI-indC*dT(ccdB)||N.A||N.A
+
-
|-
+
-
|}
+
-
<br/>
+
-
=Used T and TE Domains=
+
-
{|class="wikitable sortable"
+
-
|-
+
-
!ID!!Name (derived from)!!Description
+
-
|-
+
-
|<div id="T1">-T1</div>||indC||native T-Domain of indC from ''P. luminescens''
+
-
|-
+
-
|<div id="T2">-T2</div>||bpsA|| native T-Domain of bpsA from ''S. lavendulae''
+
-
|-
+
-
|<div id="T3">-T3</div>||entF||native T-Domain of entF from ''E. coli''
+
-
|-
+
-
|<div id="T4">-T4</div>||tycA1||native T-domain of tycA1 from ''Brevibacillus parabrevis''
+
-
|-
+
-
|<div id="T5">-T5</div>||tycC6||native T-domain of tycC6 from ''B. parabrevis''
+
-
|-
+
-
|<div id="T6">-T6</div>||delH4||native T-domain of delH4 from ''Delftia acidovorans''
+
-
|-
+
-
|<div id="T7">-T7</div>||delH5||native T-domain of delH5 from ''D. acidovorans''
+
-
|-
+
-
|<div id="T8">-T8</div>||plu2642||native T-domain of plu2642 from ''Photorhabdus luminescens''
+
-
|-
+
-
|<div id="T9">-T9</div>||plu2670||native T-domain of plu2670 from ''P. luminescens''
+
-
|-
+
-
|<div id="T10">-T10</div>||synT1||synthetic T-Domain from indC-BLAST
+
-
|-
+
-
|<div id="T11">-T11</div>||synT2||synthetic T-Domain from indC_T-Plum-BLAST
+
-
|-
+
-
|<div id="T12">-T12</div>||synT3||synthetic T-Domain from indC_T-BLAST
+
-
|-
+
-
|<div id="T13">-T13</div>||synT4||synthetic T-Domain from project_5 BLAST
+
-
|-
+
-
|<div id="T14">-T14</div>||synT5||synthetic T-Domain random 1 (indigoidine synthetases)
+
-
|-
+
-
|<div id="T15">-T15</div>||synT6||synthetic T-Domain random 2 (project_5)
+
-
|- style="border-top: double;"
+
-
|<div id="TTE1">-TTE1</div>||bpsA||native TTE-Domain of bpsA from ''S. lavendulae''
+
-
|-
+
-
|<div id="TTE2">-TTE2</div>||entF||native TTE-Domain of entF from ''E. coli''
+
-
|-
+
-
|<div id="TTE3">-TTE3</div>||tycC6||native TTE-domain of tycC6 from ''B. parabrevis''
+
-
|-
+
-
|<div id="TTE4">-TTE4</div>||delH5||native TTE-domain of delH5 from ''D. acidovorans''
+
-
|-
+
-
|}
+
-
<br/>
+
= Instruments =
= Instruments =
Line 1,485: Line 501:
= Further Recipies and Stocks=
= Further Recipies and Stocks=
-
==Acidovorans Complex Medium==
 
-
 
-
=== for 1 L ===
 
-
<html>
 
-
<ul>
 
-
<li style="font-size:14px"> 0.5 g Yeast Extract (Difco-BectonDickinson)</li>
 
-
<li style="font-size:14px"> 1.0 g Cas Amino Acids (Difco)</li>
 
-
<li style="font-size:14px"> 2.0 g Pyruvic Acid</li>
 
-
<li style="font-size:14px"> 2.0 g L-Glutamine</li>
 
-
<li style="font-size:14px"> 0.3 g KH<sub>2</sub>PO<sub>4</sub></li>
 
-
<li style="font-size:14px"> 0.3 g MgSO<sub>4</sub></li>
 
-
<li style="font-size:14px"> 2.0 g MOPS </li>
 
-
<li style="font-size:14px"> 4.0 g Chelex 100-resin (Sigma)</li>
 
-
<li style="font-size:14px"> PH adjusted to 7.2–7.3 with 5 M KOH</li>
 
-
</ul>
 
-
 
-
Important Notes:
 
-
<ul style="font-size:14px">
 
-
<li style="font-size:14px"> Fill up to 900 ml before adding pyruvic acid and L-glutamine</li>
 
-
<li style="font-size:14px"> Adjust pH</li>
 
-
<li style="font-size:14px"> Fill up to 1L</li>
 
-
<li style="font-size:14px"> Treat with Chelex for 1h</li>
 
-
<li style="font-size:14px"> Remove Chelex by filtration</li>
 
-
</ul>
 
-
</html>
 
== Ampicillin Stock Solution ==
== Ampicillin Stock Solution ==
Line 1,523: Line 514:
-
===Efficiency Test===
 
-
<html>
 
-
<div style="align:justify; font-size:14px;">
 
-
<div>
 
-
Because we doubted the effiency of our ampicillin stock solution, we prepared an effiency test. TB or LB media were prepared with different ampicillin solutions in order to detect at which concentration bacteria cells carrying a mRFP expressing plasmid with ampicillin resistence loose it.</div>
 
-
<br/>
 
-
<center>
 
-
<a class="fancybox fancyFigure" href="https://static.igem.org/mediawiki/2013/2/26/AmpTest.png" title="Figure 1: Ampicilin efficiency test.(1A) Our ampicilin stock and a different ampicilin stock from the 3rd floor was diluted in 300 µl TB media, oculated with DH10beta + pSB1A6 (FannyTest plate, 2013-08-15) and grown at 37 °C for around 1.5 day. <br/>1B: Our ampicilin stock and a different ampicilin stock from the 3rd floor was diluted in 300 µl LB media, oculated with DH10beta + pSB1A6 (FannyTest plate, 2013-08-15) and grown at 37 °C for around 1.5 day. Additionally two <b>LB media from</b> the fridge already prepared with resistence were tested.">
 
-
 
-
<img style="width:80%; padding:1%;border-style:solid;border-width:1px;border-radius: 5px;" src="https://static.igem.org/mediawiki/2013/2/26/AmpTest.png" ></img>
 
-
 
-
<figcaption style="width:80%;align:justify; font-size:14px;"><b>Figure 1A:</b> Ampicilin efficiency test: Our ampicilin stock and a different ampicilin stock from the 3rd floor was diluted in 300 µl <b>TB media</b>, oculated with DH10beta + pSB1A6 (FannyTest plate, 2013-08-15) and grown at 37 °C for around 1.5 day.<br/> <b>Figure 1B:</b> Ampicilin efficiency test: Our ampicilin stock and a different ampicilin stock from the 3rd floor was diluted in 300 µl LB media, oculated with DH10beta + pSB1A6 (FannyTest plate, 2013-08-15) and grown at 37 °C for around 1.5 day. Additionally two <b>LB media from</b> the fridge already prepared with resistence were tested.</figcaption>
 
-
</a>
 
-
</center>
 
-
<div>
 
-
<br>
 
-
<br>
 
-
<div style="align:justify; clear:both;">We can conclude that our ampicilin should still be usable, since it is at least as efficient as the ampicilin stock solution optained from a different group.</div>
 
-
</html>
 
-
 
-
== Bacitracin Stock Solution ==
 
-
{|style="font-size:14px; width:300px"
 
-
|Stock ||Bacitracin 5000 U/ml
 
-
|-
 
-
|Amount || 5 ml
 
-
|-
 
-
|Storage || -24°C freezer
 
-
|}
 
-
 
-
==<i>Brevibacillus Parabrevis</i> Glycerol Stock==
 
-
 
-
{|style="font-size:14px; width:500px; clear:both;"
 
-
|Stock || ''Brevibacillus parabrevis'' ATCC8185 glycerol stock
 
-
|-
 
-
|Amount || 16 x 1.3 ml
 
-
|-
 
-
|Storage || -80°C freezer
 
-
|}
 
== Chloramphenicol Stock Solution ==
== Chloramphenicol Stock Solution ==
Line 1,572: Line 525:
|-
|-
|Notes || Solved in 100% ethanol. Use in 1:3000 dilution
|Notes || Solved in 100% ethanol. Use in 1:3000 dilution
-
|}
 
-
 
-
== <i>D. Acidovorans</i> Glycerol Stock==
 
-
 
-
{|style="font-size:14px; width:300px; clear:both;"
 
-
|Stock || ''D. acidovorans'' glycerol stock
 
-
|-
 
-
|Amount || 10 x 1 ml
 
-
|-
 
-
|Storage || -80°C freezern
 
|}
|}
Line 1,772: Line 715:
Adjust pH to 7.0
Adjust pH to 7.0
</html>
</html>
-
 
-
==Solution for MS Preparation==
 
-
 
-
===Solution 1 (for MS Sample Preparation Neo Gen Stable Isotope Standard Kit A and B (NSK-AB, EurIsotop)===
 
-
<div style="align:justify; font-size:14px">
 
-
 
-
1. One vial of amino acid standard (NSK-A) is taken up in 1 ml methanol/water (1:1 v/v) and solubilized for ca. 15 minutes in an ultra-sonification bath. Caution: make sure that the lid is closed well and not wetted by water of the ultra-sonification bath.<br />
 
-
2. One vial of acylcarnitine standard (NSK-B) is taken up in 1 ml of methanol and solubilized for ca. 15 minutes in the ultra-sonification bath.Caution: make sure that the lid is closed well and not wetted by water of the ultra-sonification bath.<br />
 
-
Contents of both vials are transfered quantitatively with methanol in a 200 ml graduate flask.The graduate flask is filled up to the calibration mark with methanol.
 
-
Standard solutions get a consecutive number. When two 200 ml graduate flasks are prepared simultaneously, contents of both flasks can be combined, if levels of both standards (old and new) coincide. Afterwards the whole solution will be labeled with one common number.<br />
 
-
Stability of solution 1: 7 days<br /></div>
 
-
<br/>
 
-
=== Solution 2 (for MS sample preparation): butanolic hydrochloric acid (3 N), water-free ===
 
-
<div style="align:justify; font-size:14px">
 
-
1. 900 ml butanol (p.A., Merck 1.01990.1000) are cooled in an ice bath at 0°C (magnetic stirrer).
 
-
The vessel has to be closed to avoid ice condensation of water.<br/>
 
-
2. 100 ml acetylchloride (p.A., Merck 1.00031.0250) are added dropwiese from a dropping funnel over a a period of 5 minutes while stirring constantly (protection goggles and gloves!).<br/>
 
-
Caution: make sure that no water or ice gets into the solution.<br />
 
-
3. The solution is stirred for additional 10 minutes with closed lid on ice.
 
-
Stability of solution 2: 1 year <br /></div>
 
-
<br/>
 
-
=== Solution 3 (for MS sample preparation) ===
 
-
<div style="align:justify; font-size:14px">
 
-
Acetonitrile p.A./water (1:1 v/v) (83639.320DE ACETONITRIL HIPERSOLV SUPER GR REAG.PE)<br />
 
-
Stability of solution 3: 1 year<br /></div>
 

Revision as of 14:44, 16 October 2014

Contents

Kits

Kit Supplier Catalog Number
MinElute® PCR Purification Kit (250) QIAGEN 28006
Plasmid Plus Maxi Kit (25) QIAGEN 12963
Plasmid Plus Midi Kit (25) QIAGEN 12943
QIAEX II® Gel Extraction Kit (500) QIAGEN 20051
QIAGEN® Plasmid Plus Midi Kit (100) QIAGEN 12945
QIAquick® Gel Extraction Kit (250) QIAGEN 28706
QIAquick® PCR Purification Kit (250) QIAGEN 28106
QIAprep® Spin Miniprep Kit (250) QIAGEN 27106
QIAprep® Spin Miniprep Columns QIAGEN 27115
EnzChek® Ultra Xylanase Assay Kit Life Technologies E33650


Marker

Marker Supplier Catalog Number
Quick-Load® 2-Log DNA Ladder (0.1-10.0 kb) New England BioLabs N3200S
Quick-Load® 1 kb DNA Ladder New England BioLabs N0468S
50 bp DNA Ladder New England BioLabs N3236S
Gel loading solution Sigma-Aldrich Chemie GmbH G2526-5ML


Enzymes

Enzyme Supplier Catalog Number
DreamTaq Green PCR Master Mix (2X) Thermo Fisher Scientific Biosciences GmbH K1081
DreamTaq PCR MM Fermentas Life Sciences K1071
Gibson Assembly® Master Mix New England Biolabs E2611 S
Lysozyme from Chicken Egg White Sigma-Aldrich Chemie GmbH L4919-500MG
Phusion® Flash High-Fidelity PCR Master Mix Biozym Scientific GmbH F-548L
Phusion® High-Fidelity PCR Master Mix New England Biolabs M0531 L
T4 DNA Ligase New England Biolabs GmbH M0202 S
2x PCR Master mix Solution (iTaq) HISS DIAGNOSTICS GmbH 25028


Antibodies

Antibody Supplier Catalog Number
Penta·His Antibody, BSA-free QIAGEN 34660
Anti-GFP Roche 11814460001
mRFP
Goat anti Mouse IgG HRPO Dianova GmbH 115-035-003
secondary rat


Restriction Enzymes

Enzyme Supplier Catalog Number
BamI New England Biolabs R3136 S
BgIII New England Biolabs R0144 S
DpnI New England Biolabs R0176 S
EcoRI New England BioLabs R0101S
EcoRI-HF New England Biolabs R3101
HindIII-HF New England Biolabs R3104 S
KpnI-HF New England Biolabs R3142 S
MfeI-HF New England Biolabs R3589 S
NheI-HF New England BioLabs R3131 S
NotI-HF New England BioLabs R3189 S
PacI New England Biolabs R0547 S
PstI-HF New England Biolabs R3140 S
PvuI-HF New England BioLabs R3150S
SalI-HF New England Biolabs R3138 S
SpeI-HF New England BioLabs R3133 S
XbaI New England BioLabs R0145 S


Bacterial Strains

Strain Supplier Catalog Number
Delftia acidovorans DSM-39 DSMZ DSM 50251
Delftia acidovorans SPH-1 DSMZ DSM 14801
E. coli DH10ß New England Biolabs C3019
E. coli Top10 invitrogen C404010
Photorhabdus laumondii luminescens DSMZ DSM 15139
Streptomyces lavendulae lavendulae DSMZ DSM 40708
Streptomyces mobaraensis DSMZ DSM 40903


Antibiotics and Media Supplements

Antibiotic Supplier Catalog Number Concentration stock solution Dilution Solvent
Ampicillin Anhydrous Crystalline Sigma-Aldrich Chemie GmbH A9393-5G 100 mg/ml 1:1000 H2O
Ampicillin Sodium Crystalline Sigma-Aldrich Chemie GmbH A9518-5G 100 mg/ml 1:1000 H2O
Chloramphenicol Crystalline Sigma-Aldrich Chemie GmbH C0378-5G 30 mg/ml 1:3000 Ethanol
Kanamycinsulfat Mixture of Componenta Sigma-Aldrich Chemie GmbH 60615-5G 50 mg/ml 1:1000 H2O
Tetracycline Sigma-Aldrich Chemie GmbH T7660 10 mg/ml 1:1000 Ethanol
Propionic Acid Sodium Insect Cell*Culture Sigma-Aldrich Chemie GmbH P5436-100G 100mM 10mM H2O
Bacitracin Sigma-Aldrich Chemie GmbH B0125-50KU - -


Media

Medium Supplier Catalog Number
SOC Outgrowth Medium New England Biolabs GmbH B9020 S
LB BROTH BASE Th. Geyer GmbH & Co KG SA/L3022/001000
LB Broth Powder Sigma-Aldrich Chemie GmbH L3022-1KG
M9 Minimal Salts SERVA 48505.01


Buffers

Buffer Supplier Catalog Number
NEBuffer Pack #4 (green) New England Biolabs GmbH B7004 S
NEBuffer Pack #1 (yellow) New England Biolabs GmbH B7001 S
NEBuffer Pack for T4 DNA Ligase New England Biolabs GmbH B0202 S
NEBuffer Pack #2 (blue) New England Biolabs GmbH B7002 S
NEBuffer Pack #3 (red) New England Biolabs GmbH B7003 S
TAE - Buffer (50X) for Molecular Biology VWR International GmbH A4686.1000
Gel Loading Buffer Sigma-Aldrich G2526-5ML
Tris Acetate-EDTA Buffer Sigma-Aldrich T9650-1L


Other Chemicals

Chemical Supplier Catalog Number
Isopropyl B-D-Thiogalactopyranoside 1 piece Sigma-Aldrich Chemie GmbH I5502-1G
Dimethyl Sulfoxide PCR Reagent Sigma-Aldrich Chemie GmbH D9170-1VL
Glycerol Sigma Grade Sigma-Aldrich Chemie GmbH G9012-100ML
5-Bromo-4-Chloro-3-Indolyl B-D-*Galactop Sigma-Aldrich Chemie GmbH B4252-100MG
Bacteriological Agar Sigma-Aldrich Chemie GmbH A5306-250G
L-Plus-Arabinose Crystalline Sigma-Aldrich Chemie GmbH A3256-25G
Calciumchlorid Dihydrat Th. Geyer GmbH & Co KG SA/00223506/000500
Malt Extract from Starch Digestion Sigma-Aldrich Chemie GmbH M0383-100G
D(+)-Saccharose, ACS, for Micro-Biology Sigma-Aldrich Chemie GmbH 84100-1KG
Dimethyl Sulfoxide Plant Cell Culture*TE D4540-100ML
Sodium Hydroxide Anhydrous Pellets Th. Geyer GmbH & Co.KG SA/S5881/000500
TRIZMA(R) Hydrochloride PH 3.5-5.0 Sigma-Aldrich Chemie GmbH T6666-50G
L-Glutamine 200 MM Sterile Sigma-Aldrich Chemie GmbH G7513-20ML
Ethanol 96% Denatured Carl Roth GmbH & Co.KG T171.3
Propanol-2 Sigma-Aldrich Chemie GmbH 59309-1L
Natriumdodecylsulfat,SDS,99%, Ultra Pure 13904
Gold(III)-Chloride Carl Roth GmbH & Co.KG 5624.1
Pro-Leu Sigma-Aldrich Chemie GmbH P1130-1G
Nitric Acid 65% p.a. Iso Carl Roth GmbH & Co.KG X943.1
Mops, Sodium Sigma-Aldrich Chemie GmbH M9024-25G
L-Glutamine Sigma-Aldrich Chemie GmbH G7513-100ML
Chelating Resin Sigma-Aldrich Chemie GmbH C7901-50G
Potassium Hydroxide in Platellets 6751.3
Hydrochloric Acid 37% 4625.1
Pyruvic Acid Sigma-Aldrich Chemie GmbH 107360-25G
Fmoc-Orn(BOC)-OH 96.0 % 47560-5G-F
Glycerol >99.5% Sigma-Aldrich Chemie GmbH G9012-1L
Water Molecular Biology Reagent Sigma-Aldrich Chemie GmbH W4502-1L
Acetonitrile Sigma-Aldrich Chemie GmbH 34967-1L
Ascorbic Acid 99% Sigma-Aldrich Chemie GmbH A92902-100G

Electrophoresis

Reagent Supplier Catalog Number Concentration Solvent
Agarose Molecular Biology Reagent Th. Geyer GmbH & Co KG SA/A9539/000050 0.5% H2O
Agarose for Routine Use Sigma-Aldrich Chemie GmbH A9539-100G - -


Primers and Oligos

Identifier Order date Note Sequence


Plasmids

NameDateBrief DescriptionGenotypePlasmid MapGenBank-File


Instruments

Instrument Type Manufacturer
Table Centrifuge Microfuge® 18 Centrifuge Beckman CoulterTM
Table Centrifuge Microfuge® 22R Centrifuge Beckman CoulterTM
Centrifuge Allegra X-12R Beckman CoulterTM
Shaker VORTEXGENIE 2 Scientific Industries, SITM
Heating plate (magnet stirrer)MR Hei-Standard Heidolph
Heatblock QBD4 Grant
Heatblock (shakeing function) Thermomixer comfort Eppendorf
PCR-MachineMyCyclerTM thermo cyclerBioRad
PCR-MachineT100 Therma CyclerBioRad
UV-Chamber Transluminator Vilber Lourmat
Scale (fine)PioneerTM PA114C OHAUS
ScalePioneerTM PA4101C OHAUS
Fridge KTP 1750 Premium Liebherr
Freezer GP 1366 Premium Liebherr
HoodTischabzugWesemann® Laboreinrichtung
Draw-Off PumpVacuhand controlVacubrand
Incubator HT Multitron Version 2 INFORS
Plate Reader
Computer Sun Microsystems
UV/VIS Spectrometer Ultrospec 3300 pro Amersham Biosciences
Photometer NanoDrop® ND-1000 Spectrophotometer peQLab Biotechnologie GmbH
Photometer NanoVue General Electric
Ice Machine MF22SCOTSMAN®
Gel Electrophoresis Chamber Mupid®-One Advance
Cell Density Meter Ultrospec10 Amersham Biosciences
PH-Meter PH-Meter 765 Calimatic Knick
Incubator Heraeus Thermo Scientific
Lyophilizer Gefriertrocknungsanlage ALPHA 1-2 LD Plus Best. Nr. 101521 Christ
Ultra Sonification Stick Sonoplus Gm 2070 (2002)Bandelin
Vacuum Manifold Qiavac 24 Plus QIAGEN
Gas Cartridge Ventil CV470 Burner 13883
NuPAGE(R) Novex 3-8% Tris-Acetate Gel 1 SDS-Gel Life Technologies GmbH


Mass Spectrometry and Neonate Screening

Instrument Type Manufacturer Order number
Micro Filter Plates Multiscreen 96 well Merck Millipore MSHVN4550
Microtiter Plates 96 well Mikrotiter plates made of Polypropylen (F-Form) Greiner 655201
Microtiter Plates Locks Micromats
Aluminium Foil
Mass Spectrometer Quattro Ultima (ESI-MS/MS) Micromass (today: Waters)
Methanol (20837.320 METHANOL HIPERSOLV HPLC ISOCRATIC GRADE)


Lab Materials

Material Name Manufacturer
Bottle 50 ml, 100 ml, 200 ml, 500 ml, 1000 ml
Beaker 50 ml, 100 ml, 200 ml, 500 ml 1000 ml
Conical Flask 500 ml, 1000 ml, 100 ml
Conical Flask 300 ml
Single Channel Pipette Pipetman® P2, P20, P200, P1000 Gilson
Multichannel Pipette
Multistep Pipette
Pipette
0.2 ml PCR Tube, Flat Cap, Natural I1402-8200 Starlab GmbH
Microcentrifuge Tubes
Centrifugetubes
PCR 8-Strips
Gloves Nitril Gr. L Starlab powderfree Starlab
Gloves Nitril Gr. M Starlab powderfree Starlab
Gloves Nitril Gr. S Starlab powderfree Starlab
Filtertips
Inoculating Loop
Disposal Bags
Flasks
Plate
Petri Dishes P5731-500EA Sigma-Aldrich Chemie GmbH
Petri Dishes N221.2 Carl Roth GmbH & Co.KG
TipOne® Pipette Tip S1110-1700 Starlab GmbH
TipOne® Pipette Tip, 1000μl, Graduated S1111-2721 Starlab GmbH
NeoBox-81 6er Set, je 1 x Transparent, g 22916
NeoLab-Marker for Reaction-Flaks 19079
Gene Pulser/MicroPulser Cuvettes, 0.1 cm 165-2089
NeoLab-Paper Scissors, 23 cm Long, Even 23272
TipOne® Pipette Tip, 200μl S1110-1700 Starlab GmbH
TipOne® Pipette Tip, 10μl, Graduated, Re S1111-3700 Starlab GmbH
Pipette, 5 ml 606180 Greiner bio-one GmbH
Ring out of Plumbum with Vinyl Coating, 57 mm In 310161013 NEOLAB GmbH
TipOne® Pipette Tip 10μl, Graduated, Rac S1111-3800 Starlab GmbH
Reaction Tube,S.L.1.5 ml,Colorless. 12682 Eppendorf, Fisher Scientific GmbH
Reaction Tube,S.L.,2 ml, Colorless 12776 Eppendorf
NeoTape-Writing Tape, 13 mm, Gray 280126114 NEOLAB GMBH
NeoTape-Writing Tape, 25 mm, Salmon-Colored 280126229 NEOLAB GMBH
10 ml Serological Pipette, Filter, Sterile E4860-1011 Starlab GmbH
Gloves Latex + Alovera L 14089
Gloves Latex + Alovera M 14088
Gloves Latex + Alovera S 14087
PH-Stripes WHAT10362000 PANPEHA PLUS
100 Run24Barcode 20110099-100 GATC Biotech AG
Tube Conical, Polypropylen, 50 ml 352070 NEOLAB GMBH
Gene Pulser/MicroPulser Cuvettes, 0.1 cm 165-2089 Bio-Rad Laboratories GmbH
0.2 ml 8-Tube Strips Without Caps, Nature TBS-0201 Bio-Rad Laboratories GmbH
Round-Bottom Tubule, Polypropylen, 14 ml 352059 NEOLAB GMBH
Optical Flat 8-Cap Strips TCS-0803 Bio-Rad Laboratories GmbH
Inoculation Loop 10 μl, Blue, Sterile 2900254437
Corning Serological Pipette 50 ml 14303
Weighing Dish 500 ST 1884.1 Carl Roth GmbH & Co.KG
Filter Paper Z274836-1PAK Sigma-Aldrich Chemie GmbH
Inoculation Loop 10 µl 2900254437 NEOLAB GMBH


Further Recipies and Stocks

Ampicillin Stock Solution

Stock Ampicillin 100 mg / ml
Amount 50 ml
Storage -24°C freezer
Notes Use in 1:1000 dilution


Chloramphenicol Stock Solution

Stock Chloramphenicol 30 mg / ml
Amount 10 ml
Storage -24°C freezer
Notes Solved in 100% ethanol. Use in 1:3000 dilution

E.Coli BAP 1 Glycerol Stock

Stock E. coli BAP1 glycerol stock
Amount 2 x 1.3 ml
Storage -80°C freezer

E.Coli BAP1, Competent

Stock E. coli BAP1
Amount 18 x 100 µl
Storage -80°C freezer
Notes Grows extremely fast. Be careful with miniPreps,
at least in cultures with ampicillin it tends
to degrade all available ampicillin
and then lose the respective plasmid.

E. coli BAP1-pKD46 Glycerol Stock

What E. coli BAP1-pKD46 glycerol stock (Ampr)
Amount 2 x 1.3 ml
Storage -80°C freezer
Notes Grow at 30°C only! Growth at 37°C
will lead to loss of pKD46 plasmid.

E.Coli BAP1-pLF03 Glycerol Stock

Stock E. coli BAP1-pLF03 glycerol stock (Ampr)
Amount 1 x 1.3 ml
Storage -80°C freezer
Notes Might have low amount of plasmid-carrying bacteria
due to long culturing time
(all Amp in medium cleaved)

E. Coli DH5α-pCP20 Glycerol Stock

Stock E. coli DH5α-pCP20 glycerol stock (Ampr, Cmr)
Amount 2 x 1.3 ml
Storage -80°C freezer
Notes Grow at 30°C only!
Growth at 37°C will lead to loss of pCP20 plasmid.

Heat-Shock Competent E. Coli TOP10

Stock E. coli TOP10, Heat-shock competent
Amount 200 x 100 µl
Storage -80°C freezer
Notes Verified on 2013-06-07.

Figure 3 Control transformation of competent TOP10 cells with 80 ng pSB4K5 with insert J04450 (IPTG-inducible mRFP production). Left: transformation with plasmid; right: transformation with water.

E. Coli TOP10-BBa I746200/pSB1AK3 Glycerol Stock

Stock E. coli TOP10-(BBa I746200 in pSB1AK3) (Ampr, Kanr)
Amount 1 x 1.3 ml
Storage -80°C freezer

E. Coli TOP10-BBa J04450/pSB3C5 Glycerol Stock

Stock E. coli TOP10-(BBa_J04450 in pSB3C5) glycerol stock (Cmr)
Amount 1 x 1.3 ml
Storage -80°C freezer

E. Coli TOP10-pIK1 Glycerol Stock

Stock E. coli TOP10-pIK1 glycerol stock (Cmr)
Amount 1 x 1.3 ml
Storage -80°C freezer

E. Coli TOP10-pKD46 Glycerol Stock

Stock E. coli TOP10-pKD46 glycerol stock (Ampr)
Amount 2 x 1.3 ml
Storage -80°C freezer
Notes Grow at 30°C only! Growth at 37°C
will lead to loss of pKD46 plasmid.

E. Coli TOP10-pMM65 Glycerol Stock

Stock E. coli TOP10-pMM65 glycerol stock (Kanr)
Amount 1 x 1.3 ml
Storage -80°C freezer

IPTG Stock Solution

Stock IPTG 23.8 mg / ml
Amount 10 ml
Storage -20°C freezer
Notes Use 0.1 to 1 mM

Kanamycin Stock Solution

Stock Kanamycin 50 mg / ml
Amount 10 ml
Storage -24°C freezer
Notes Use in 1:1000 dilution

M9 Medium

Name M9 Minimal Salts 5x, Powder
Amount 1 l
Storage room temperature
Notes close tightly, hygroscopic

  • Add 200 ml of sterile M9 salt solution to 750 ml sterile, distilled H2O (45-50°C)
  • Add sterile 20 ml 20% Glucose-solution, 2 ml 1 M MgSO4 and (optionally) 1 M CaCal

Reactivation Medium

for 1L

  • 5.0 g Peptone
  • 3.0 g Meat extract
  • 1000.0 ml Distilled water
Adjust pH to 7.0

Retrieved from "http://2014.igem.org/Http://2014.igem.org/Team:Heidelberg/pages/Notebook/Materials"