"
Team:Bielefeld-CeBiTec/Notebook/Primer
From 2014.igem.org
Revision as of 13:12, 27 August 2014 by SebastianBlunk (Talk | contribs)
Primerlist
| Primer ID | Primer sequence |
|---|---|
| General | |
| VF-Primer (20 bp) | 5‘-CCACCTGACGTCTAAGAAAC-3‘ |
| VR-Primer (22 bp) | 5‘-GTATTACCGCCTTTGAGTGAGC-3‘ |
| Module 1 | |
| Gene synthesis of the fumarate reductase (fumA, fumBCD) of B. succinogenes | |
| pSB1C3_pre_Fum_A (43 bp) | 5‘-GTCTGCATTTTCTCCTCTTTCTCTAGAAGCGGCCGCGAATTCC-3‘ |
| pSB1C3_suf_Fum_A (43 bp) | 5‘-CCAAAGCGCCGAATGCGTAATACTAGTAGCGGCCGCTGCAGTC-3‘ |
| pSB1C3_pre_Fum_BCD (42 bp) | 5‘-TTGGTCATTTTCTCCTCTTTCTCTAGAAGCGGCCGCGAATCC-3‘ |
| pSB1C3_suf_Fum_BCD (43 bp) | 5‘-GGGGTGTGGCGTCGATCTAATACTAGTAGCGGCCGCTGCAGTC-3‘ |
| FumBCD_fwd (24 bp) | 5‘-CGGCCGCTTCTAGAGAAAGAGGAG-3‘ |
| FumBCD_rev (22 bp) | 5‘-CGGCCGCTACTAGTATTAGATC-3‘ |
| FumA_seq (20 bp) | 5‘-TGGAAGGCACCTTGGTTCAG-3‘ |
| FumB_seq (20 bp) | 5‘-GCTCTGACCATGGCACATCG-3‘ |
| Overexpression of the fumarate reductase (frd) of E. coli | |
| T7_frd_Ec_fwd (64 bp) | 5‘-TAATACGACTCACTATAGGGAAAGAGGAGAAATACTAGATGCAAACCTTTCAAGCCGATCTTGC-3‘ |
| T7_frd_Ec_rev (44 bp) | 5‘-TGCAGCGGCCGCTACTAGTATTAGATTGTAACGACACCAATCAG-3‘ |
| pSB1C3_frd_pre (45 bp) | 5‘-GCAAGATCGGCTTGAAAGGTTTGCATCTAGTATTTCTCCTCTTTC-3‘ |
| pSB1C3_frd_suf (47 bp) | 5‘-CTGATTGGTGTCGTTACAATCTAATACTAGTAGCGGCCGCTGCAGTC-3‘ |
| Replacement of the succinate carrier (dcuB) by porines (oprF) in the genome | |
| porine_fwd_FRT (73 bp) | 5‘-TTATAAGAACCCGTACATCGCGGCGAAGATCCAGCCGAAGACGCACGATATTGACAGCTAGCTCAGTCCTAGG-3‘ |
| porine_rev_FRT (46 bp) | 5‘-CGCCCTTTAGTGAGGGTTAATTTTACTTAGCTTGGGCTTCAACCT-3‘ |
| pR6K_dcuB_fwd (46 bp) | 5‘-AGGTTGAAGCCCAAGCTAAGTAAAATTAACCCTCACTAAAGGGCG-3‘ |
| pR6K_dcuB_rev (73 bp) | 5‘-ATGTTATTTACTATCCAACTTATCATAATACTGATATGTCTGTTTTATGGTAATACGACTCACTATAGGGCTC-3‘ |
| dcuB_del_kon1 (20 bp) | 5‘-CGCCGCATTCTGTGAATAGC-3‘ |
| dcuB_del_kon2 (20 bp) | 5‘-CCCTGGCACAAGTTACATCC-3‘ |
| pR6K_dcuB_short (22 bp) | 5‘-AATTAACCCTCACTAAAGGGCG-3‘ |
| NAPH/NAP+ sensor (soxRS) | |
| soxR_fwd (64 bp) | 5‘-TTGACAGCTAGCTCAGTCCTAGGTATTGTGCAAAGAGGAGAAAATGGAAAAGAAATTACCCCGC-3‘ |
| soxR_rev (37 bp) | 5‘-GCTTCTAGAGTTAGTTTTGTTCATCTTCCAGCAAGCG-3‘ |
| pSB1C3_suf_soxR (54 bp) | 5‘-TAGGACTGAGCTAGCTGTCAATACTAGTAGCTACTAGTAGCGGCCGCTGCAGTC-3‘ |
| pSB1C3_pre_soxR (43 bp) | 5‘-TGGAAGATGAACAAAACTAACTCTAGAAGCGGCCGCGAATTCC-3‘ |
| soxS_GFP_fwd (74 bp) | 5‘-TTATACTCCCCAACAGATGAATTAACGAACTGAACACTGAAAAGAGGCAGATTTATGCGTAAAGGAGAAGAACT-3‘ |
| soxS_GFP_rev (42 bp) | 5‘-TGCAGCGGCCGCTACTAGTATATAAACGCAGAAAGGCCCACC-3‘ |
| pSB1C3_pre_soxS (78 bp) | 5‘-TAATTCATCTGTTGGGGAGTATAATTCCTCAAGTTAACTTGAGGTAAAGCGATTTCTCTAGAAGCGGCCGCGAATTCC-3‘ |
| pSB1C3_suf_soxS (43 bp) | 5‘-TGGGCCTTTCTGCGTTTATATACTAGTAGCGGCCGCTGCAGTC-3‘ |
| Expression of the ccm cluster and cytochrome C (pccH) | |
| ccm_fwd (58 bp) | 5‘-CTCAGTCCTAGGTATTGTGCAGAGAAAGAGGAGAAAATGCTTGAAGCCAGAGAGTTAC-3‘ |
| ccm_rev (47 bp) | 5‘-TGCAGCGGCCGCTACTAGTATTATTTACTCTCCTGCGGCGACAAATG-3‘ |
| pSB1C3_suf_ccm (50 bp) | 5‘-CATTTGTCGCCGCAGGAGAGTAAATAATACTAGTAGCGGCCGCTGCAGTC-3‘ |
| pSB1C3_pre_ccm (28 bp) | 5‘-CTCTGCTAGCACAATACCTAGGACTGAG-3‘ |
| pccH_fwd (55 bp) | 5‘-ATTCGCGGCCGCTTCTAGAGAAAGAGGAGAAAATGCAAAAAACAACTATTCTTTC-3‘ |
| pccH_rev (40 bp) | 5‘-TGCAGCGGCCGCTACTAGTATCAGTACGTGACCGTGATGG-3‘ |
| pSB1C3_pre_pccH (58 bp) | 5‘-GAAAGAATAGTTGTTTTTTGCATTTTCTCCTCTTTCTCTAGAAGCGGCCGCGAATTCC-3‘ |
| pSB1C3_suf_pccH (43 bp) | 5‘-CCATCACGGTCACGTACTGATACTAGTAGCGGCCGCTGCAGTC-3‘ |
| Module 2 | |
| Gene synthesis of the RuBisCO of Synechoccus elangatus (rbcLS) and Halothiobacillus neapolitanus (ccbLS) and the phosphoribulokinase A (prkA) of Synechoccus elangatus | |
| pSB1C3_pre_rbcLS (44 bp) | 5‘-CATCTAGTATTTCTCCTCTTTCTCTAGAAGCGGCCGCGAATTCC-3‘ |
| pSB1C3_suf_rbcLS (43 bp) | 5‘-ATCGTCCGGGCCGCTACTAA-TACTAGTAGCGGCCGCTGCAGTC-3‘ |
| pSB1C3_pre_ccbLS (44 bp) | 5‘-CATCTAGCGGCCGCTACTAGTATTAGATCTATTTCTCCTCTTTCTCTAGAAGCGGCCGCGAATTCC-3‘ |
| pSB1C3_suf_ccbLS (43 bp) | 5‘-TGGTTTACCGCGGCAACTAATACTAGTAGCGGCCGCTGCAGTC-3‘ |
| pSB1C3_pre_prkA (43 bp) | 5‘-ACACGATCTGGCTTGCTCATCTCTAGAAGCGGCCGCGAATTCC-3‘ |
| pSB1C3_suf_prkA (43 bp) | 5‘-CGGTTGCCGCTAGCGTTTAATACTAGTAGCGGCCGCTGCAGTC-3‘ |
| Hneap_seq (20 bp) | 5‘-TACAGTTTGGACCGACCTTC-3‘ |
| Selan_seq (20 bp) | 5‘-GTACATGGACGACTGTATGG-3‘ |
| Insert amplification of the gene synthesis of the RuBisCO of S. elangatus (SELAN) | |
| Selan_fwd (20 bp) | 5‘-GGCCGCTTCTAGAGAAAGAG-3‘ |
| Selan_rev (20 bp) | 5‘-GCGGCCGCTACTAGTATTAG-3‘ |
| Sequencing primer for the addgene plasmid pHNCBcsoS1D | |
| add_seq1 (20 bp) | 5‘-AGCGTGTCGTAGTGCGTATC-3‘ |
| add_seq2 (20 bp) | 5‘-ACCACTTCTGGCTGCACAAC-3‘ |
| add_seq3 (20 bp) | 5‘-CTGGCAACCGGCTTGATTAC-3‘ |
| add_seq4 (20 bp) | 5‘-AAATCACCCACGCCAAACCG-3‘ |
| add_seq5 (20 bp) | 5‘-GATGCGATTGGCTGCATACC-3‘ |
| add_seq6 (20 bp) | 5‘-GCCGTAATATCCAGCTGAAC-3‘ |
| Integration of the prkA in the plasdmid pHnCBcsoS1D | |
| pHnCBcsoS1D_fwd (44 bp) | 5‘-CGGTTGCCGCTAGCGTTTAAGTGAGCGGATAACAATTTCAG-3‘ |
| pHNCBcsoS1D_rev (45 bp) | 5‘-CATCTAGTATTTCTCCTCTTTTCCACACATTATACGAGCCGGAAG-3‘ |
| prkA_pHn_rev (47 bp) | 5‘-CTGAAATTGTTATCCGCTCACTTAAACGCTAGCGGCAACCGGTGCTG-3‘ |
| prkA_pHn_fwd (53 bp) | 5‘-CTTCCGGCTCGTATAATGTGTGGAAAAGAGGAGAAATACTAGATGAGCAAGCC-3‘ |
| Shell associated protein of the carboxysome(sap respectively csoS2) | |
| csoS2_sap_1_fwd (43) | 5‘-ATTCGCGGCCGCTTCTAGAGGTCAAGTCTGTCATTGCGCTGTG-3‘ |
| pSB1C3_pre_sap_1 (43 bp) | 5‘-AGCGCAATGACAGACTTGACCTCTAGAAGCGGCCGCGAATTCC-3‘ |
| csoS2_sap_1_rev (38 bp) | 5‘-GCGCCGAACATGACTTTATCCGCTTGTGAAGGCTTTGC-3‘ |
| sap_2_fwd (22 bp) | 5‘-AAGTCATGTTCGGCGCCACAGC-3‘ |
| sap_2_rev (20 bp) | 5‘-GCCGCTACTAGTATTAACCG-3‘ |
| pSB1C3_suf_sap2 (40 bp) | 5‘-CCGGCGGCGCGCGCGGTTAATACTAGTAGCGGCCGCTGCA-3‘ |
| seq_sap_2 (20 bp) | 5‘-TGCCCGTCACCGGTAATGAG-3‘ |
| Carbonic anhydrase of the carboxysome (can) | |
| csoS3_can_fwd (50 bp) | 5‘-ATTCGCGGCCGCTTCTAGAGTTAAGTAAAGTGTAACGATCATGAACACCC-3‘ |
| pSB1C3_pre_can (46 bp) | 5‘-GTTCATGATCGTTACACTTTACTTAACTCTAGAAGCGGCCGCGAAT-3‘ |
| csoS3_can_rev (41 bp) | 5‘-TGCAGCGGCCGCTACTAGTATCATGCGGATGCAACCTCTTC-3‘ |
| pSB1C3_suf_can (46 bp) | 5‘-TCGAAGAGGTTGCATCCGCATGATACTAGTAGCGGCCGCTGCAGTC-3‘ |
| Shell proteins of the carboxysome (csoS4AB and csoS1CAB) | |
| fw_pSB1C3_csoS4A (47 bp) | 5‘-ATTCGCGGCCGCTTCTAGAGAAATCATGCAAGTTGAGAAAACG-3‘ |
| rv_csoS4A_PstI (43 bp) | 5‘-GAGGGATAAGACTTGCTTCCGGCAGCCTCTCGTGCTGCCGATG-3‘ |
| fw_PstI_csoS4A (51 bp) | 5‘-TCATCGGCAGCACGAGAGGCTGCCGGAAGCAAGTCTTATCCCTCTGATTTG-3‘ |
| rv_SpeI_Intergen (52 bp) | 5‘-GTCGTTGCCATAACAGATCCTCTAAAACTAGGTTGAGCACGTTCAGAACGAG-3‘ |
| fw_SpeI_Intergen (49 bp) | 5‘-TCGTTCTGAACGTGCTCAACCTAGTTTTAGAGGATCTGTTATGGCAACG-3‘ |
| rv_csoS1_pSB1C3 (50 bp) | 5‘-CGGACTGCAGCGGCCGCTACTAGTATTAGCTATTCAGATTTGCGATACAC-3‘ |
| fw_csoS1_pSB1C3 (46 bp) | 5‘-GTGTATCGCAAATCTGAATAGCTAATACTAGTAGCGGCCGCTGCAG-3‘ |
| rv_pSB1C3_csoS4A (47 bp) | 5‘-AACGTTTTCTCAACTTGCATGATTTCTCTAGAAGCGGCCGCGAATTC-3‘ |
| Pore protein of the carboxysome (csoS1D) | |
| csoS1D_fwd (40 bp) | 5‘-ATTCGCGGCCGCTTCTAGAGTGTCTAGTAGGGAAGATGCG-3‘ |
| csoS1D_rev (41 bp) | 5‘-TGCAGCGGCCGCTACTAGTATTAGAACCCTTCAGCGCGACG-3‘ |
| pSB1C3_suf_csoS1D (43bp) | 5‘-GTCGCGCTGAAGGGTTCTAATACTAGTAGCGGCCGCTGCAGTC-3‘ |
| pSB1C3_pre_csoS1D (43 bp) | 5‘-CGCATCTTCCCTACTAGACACTCTAGAAGCGGCCGCGAATTCC-3‘ |
| GFP fusion with the shell proteins and the shell-associated protein | |
| fw-csoS1A-GFP (45 bp) | 5‘-CTGCCTAAGGCGCCACAAGCCATGCGTAAAGGAGAAGAACTTTTC-3‘ |
| rv-csoS1B-GFP (55 bp) | 5‘-GTGAGCCGTCTTAGGAATATCTGACTTATTATTTGTATAGTTCATCCATGCCATG-3‘ |
| fw-GFP-csoS1B (48 bp) | 5‘-CATGGATGAACTATACAAATAATAAGTCAGATATTCCTAAGACGGCTC-3‘ |
| rv-GFP-csoS1A (45 bp) | 5‘-GAAAAGTTCTTCTCCTTTACGCATGGCTTGTGGCGCCTTAGGCAG-3‘ |
| Gluco switch using sRNA | |
| pSB1C3_pre_sRNApfkA (43 bp) | 5‘-TGTGGCTAAAAGAATTAGTACTCTAGAAGCGGCCGCGAATTCC-3‘ |
| pSB1C3_suf_sRNApfkA (43 bp) | 5‘-TGGGCCTTTCTGCGTTTATATACTAGTAGCGGCCGCTGCAGTC-3‘ |
| sRNA:pfkB_fwd (46 bp) | 5‘-TACCATGCATAGATATGCAACTGTTTTCTGTTGGGCCATTGCATTG-3‘ |
| sRNA:pfkB_rev (47 bp) | 5‘-ACAGTTGCATATCTATGCATGGTACTCTAGAAGCGGCCGCGAATTCC-3‘ |
| Amplification of purification vector with T7, RBS and intein tag with chitin binding domain | |
| int_RBS_fw (1) (42 bp) | 5‘-CTATAGGGAAAGAGGAGAAATGCATCACGGGAGATGCGCTAG-3‘ |
| RBS_int_rev (2) (44 bp) | 5‘-CTAGCGCATCTCCCGTGATGCATTTCTCCTCTTTCCCTATAGTG-3‘ |
| RBS_rev (3) (22 bp) | 5‘-TTTCTCCTCTTTCCCTATAGTG-3‘ |
| int_fw (4) (22 bp) | 5‘-TGCATCACGGGAGATGCACTAG-3‘ |
| int_pSB1C3_rev (5) (50 bp) | 5‘-GACTGCAGCGGCCGCTACTAGTATCATTGAAGCTGCCACAAGGCAGGAAC-3‘ |
| pSB1C3_int_fw (6) (46 bp) | 5‘-CTGCCTTGTGGCAGCTTCAATGATACTAGTAGCGGCCGCTGCAGTC-3‘ |
| Expression of the SBPase (glpX)for the calvin cycle and for purification | |
| fw-pSB1C3-BBa_B0034-SBPase (58 bp) | 5‘-GAATTCGCGGCCGCTTCTAGAGAAAGAGGAGAAAATGAGGGAATTGAAAAGCGAAAAG-3‘ |
| rv-pSB1C3-SBPase (56 bp) | 5‘-GACTGCAGCGGCCGCTACTAGTATCAATGATAAGCTTCAATAAATTGGTATTTTAC-3‘ |
| fw-SBPase-Schnittstelle (56 bp) | 5‘-AAAAGCAAAAGGCTGCGTAAATTTGGACGCATCTTTAACAGAAAATATGAAATCAG-3‘ |
| rv-SBPase-Schnittstelle (46 bp) | 5‘-TTTTCTGTTAAAGATGCGTCCAAATTTACGCAGCCTTTTGCTTTTG-3‘ |
| fw-SBPase-pSB1C3 (57 bp) | 5‘-AATTTATTGAAGCTTATCATTGATACTAGTAGCGGCCGCTGCAGTCCGGCAAAAAAG-3‘ |
| rv-SBPase-BBa_B0034-pSB1C3 (65 bp) | 5‘-TTTTCGCTTTTCAATTCCCTCATTTTCTCCTCTTTCTCTAGAAGCGGCCGCGAATTCCAGAAATC-3‘ |
| SBPase_purify_fw (45 bp) | 5‘-CTATAGGGAAAGAGGAGAAATATGAGGGAATTGAAAAGCGAAAAG-3‘ |
| SBPase_purify_rev (53 bp) | 5‘-CTAGTGCATCTCCCGTGATGCACGTAAAATACCAATTTATTGAAGCTTATCAT-3‘ |
| fw_pSB1_SBPase (58 bp) | 5‘-GAATTCGCGGCCGCTTCTAGAGAAAGAGGAGAAAATGAGGGAATTGAAAAGCGAAAAG-3‘ |
| rv_pSB1_SBPase (56 bp) | 5‘-GACTGCAGCGGCCGCTACTAGTATCAATGATAAGCTTCAATAAATTGGTATTTTAC-3‘ |
| fw_SBPase_XbaI (56 bp) | 5‘-AAAAGCAAAAGGCTGCGTAAATTTGGACGCATCTTTAACAGAAAATATGAAATCAG-3‘ |
| rv_SBPase_XbaI (46 bp) | 5‘-TTTTCTGTTAAAGATGCGTCCAAATTTACGCAGCCTTTTGCTTTTG-3‘ |
| fw_SBPase_pSB1C3 (57 bp) | 5‘-AATTTATTGAAGCTTATCATTGATACTAGTAGCGGCCGCTGCAGTCCGGCAAAAAAG-3‘ |
| rv_SBPase_pSB1 (65 bp) | 5‘-TTTTCGCTTTTCAATTCCCTCATTTTCTCCTCTTTCTCTAGAAGCGGCCGCGAATTCCAGAAATC-3‘ |
| Amplification of E. coli fructose bisphosphate aldolase II (fbaA) for purification | |
| fbaA_purify_fw (50 bp) | 5‘-CTATAGGGAAAGAGGAGAAATATGTCTAAGATTTTTGATTTCGTAAAACC-3‘ |
| fbaA_purify_rev (41 bp) | 5‘-CTAGTGCATCTCCCGTGATGCACAGAACGTCGATCGCGTTC-3‘ |
| fbaA_PstI_fw (42 bp) | 5‘-CAAAGCGAACGAAGCTTATCTGCAAGGTCAGCTGGGTAACCC-3‘ |
| fbaA_PstI_rev (43 bp) | 5‘-CTTTCGGGTTACCCAGCTGACCTTGCAGATAAGCTTCGTTCGC-3‘ |
| Amplification of E. coli transketolase I (tktA) for purification | |
| tktA_purify_fw (43 bp) | 5‘-CTATAGGGAAAGAGGAGAAATATGTCCTCACGTAAAGAGCTTG-3‘ |
| tktA_purify_rev (43 bp) | 5‘-CTAGTGCATCTCCCGTGATGCACAGCAGTTCTTTTGCTTTCGC-3‘ |
| tktA_PstI_fw (41 bp) | 5‘-GTTCGGTCCGCTGTTGCCGGAACTCCTCGGCGGTTCTGCTG-3‘ |
| tktA_PstI_rev (42 bp) | 5‘-CAGGTCAGCAGAACCGCCGAGGAGTTCCGGCAACAGCGGACC-3‘ |
| Amplification of C. glutamicum phosphoglycerate kinase (pgk) for purification | |
| cg_pgk_purify_fw (42 bp) | 5‘-CTATAGGGAAAGAGGAGAAATATGGCTGTTAAGACCCTCAAG-3‘ |
| cg_pgk_purify_rev (42 bp) | 5‘-CTAGTGCATCTCCCGTGATGCACTGAGCGAGAATTGCAACGC-3‘ |
| Amplification of E. coli glyceraldehyde 3-phosphate dehydrogenase (gapA) for purification | |
| gapA_purify_fw (47 bp) | 5‘-CTATAGGGAAAGAGGAGAAATATGACTATCAAAGTAGGTATCAACGG-3‘ |
| gapA_purify_rev (43 bp) | 5‘-CTAGTGCATCTCCCGTGATGCATTTGGAGATGTGAGCGATCAG-3‘ |
| gapA_NotI_fw (44 bp) | 5‘-CTCACAAAGACTGGCGCGGCGGCCGTGGCGCTTCCCAGAACATC-3‘ |
| gapA_NotI_rev (43 bp) | 5‘-GATGATGTTCTGGGAAGCGCCACGGCCGCCGCGCCAGTCTTTG-3‘ |
| Amplification of S. elongatus phosphoribulokinase (prkA) for purification | |
| prkA_purify_fw (40 bp) | 5‘-CTATAGGGAAAGAGGAGAAATATGAGCAAGCCAGATCGTG-3‘ |
| prkA_purify_rev (42 bp) | 5‘-CTAGTGCATCTCCCGTGATGCAAACGCTAGCGGCAACCGGTG-3‘ |
| Amplification of E. coli pyruvate kinase (pykF) for purification | |
| pykF_purify_fw (47 bp) | 5‘-CTATAGGGAAAGAGGAGAAATATGAAAAAGACCAAAATTGTTTGCAC-3‘ |
| pykF_purify_rev (42 bp) | 5‘-CTAGTGCATCTCCCGTGATGCACAGGACGTGAACAGATGCGG-3‘ |
| pykF_PstI_fw (48 bp) | 5‘-ACCGTCTGGGTAAAGAACTGGCTCTGCAAAGCGGTCTGGCACACAAAG-3‘ |
| pykF_PstI_rev (43 bp) | 5‘-CACCTTTGTGTGCCAGACCGCTTTGCAGAGCCAGTTCTTTACC-3‘ |
| Amplification of E. coli D-lactate dehydrogenase (ldhA) for purification | |
| ldhA_purify_fw (44 bp) | 5‘-CTATAGGGAAAGAGGAGAAATATGAAACTCGCCGTTTATAGCAC-3‘ |
| ldhA_purify_rev (42 bp) | 5‘-CTAGTGCATCTCCCGTGATGCAAACCAGTTCGTTCGGGCAGG-3‘ |
| Amplification of E. coli glucokinase (glk) for purification | |
| glk_purify_fw (44 bp) | 5‘-CTATAGGGAAAGAGGAGAAATATGACAAAGTATGCATTAGTCGG-3‘ |
| glk_purify_rev (43 bp) | 5‘-CTAGTGCATCTCCCGTGATGCACAGAATGTGACCTAAGGTCTG-3‘ |
| Amplification of E. coli glucose 6-phosphate-1-dehydrogenase (zwf) for purification | |
| zwf_purify_fw (41 bp) | 5‘-CTATAGGGAAAGAGGAGAAATATGGCGGTAACGCAAACAGC-3‘ |
| zwf_purify_rev (45 bp) | 5‘-CTAGTGCATCTCCCGTGATGCACTCAAACTCATTCCAGGAACGAC-3‘ |
| Module 3 | |
| Backbone for the isobutanol plasmid | |
| fw_kivD_pSB1C3 (53 bp) | 5‘-GCAAACTATTTGCTGAACAAAATAAATCATAATACTAGTAGCGGCCGCTGCAG-3‘ |
| rv_alsS_pSB1C3 (65 bp) | 5‘-TTTTTGTTCTTTTGTTGCTTTTGTCAACATTTTCTCCTCTTTCTCTAGAAGCGGCCGCGAATTC-3‘ |
| fw_adhA_pSB1C3 (51 bp) | 5‘-GGCCGAATGGTCATTGATTTTACTAAATAATACTAGTAGCGGCCGCTGCAG-3‘ |
| fw_adhA_pSB1C3 (42 bp) | 5‘-CATTGATTTTACTAAATAATACTAGTAGCGGCCGCTGCAGTC-3‘ |
| alsS | |
| fw_pSB1C3_alsS (79 bp) | 5‘-TTCGCTAAGGATGATTTCTGGAATTCGCGGCCGCTTCTAGAGAAAGAGGAGAAAATGTTGACAAAAGCAACAAAAGAAC-3‘ |
| rv_ilvC_alsS (64 bp) | 5‘-GATTCAGTGTATTGAAGTAGTTAGCCATTTTCTCCTCTTTCTAGAGAGCTTTCGTTTTCATGAG-3‘ |
| ilvC | |
| fw_alsS_ilvC (65 bp) | 5‘-CTCATGAAAACGAAAGCTCTCTAGAAAGAGGAGAAAATGGCTAACTACTTCAATACACTGAATC-3‘ |
| rev_ilvD_ilvC (56 bp) | 5‘-GTGGCGGAACGGTACTTAGGCATTTTCTCCTCTTTTTAACCCGCAACAGCAATACG-3‘ |
| ilvD | |
| fw_ilvC_ilvD (57 bp) | 5‘-CGTATTGCTGTTGCGGGTTAAAAAGAGGAGAAAATGCCTAAGTACCGTTCCGCCAC-3‘ |
| rv_kivD_ilvD (67 bp) | 5‘-GTCTAATAGGTAATCTCCTACTGTATACATTTTCTCCTCTTTTTAACCCCCCAGTTTCGATTTATC-3‘ |
| kivD | |
| fw_ilvD_kivD (67 bp) | 5‘-GATAAATCGAAACTGGGGGGTTAAAAAGAGGAGAAAATGTATACAGTAGGAGATTACCTATTAGAC-3‘ |
| rv_pSB1C3_kivD (74 bp) | 5‘-ACCTTGCCCTTTTTTGCCGGACTGCAGCGGCCGCTACTAGTATTATGATTTATTTTGTTCAGCAAATAGTTTGC-3‘ |
| rv_adhA_kivD (69 bp) | 5‘-GTGTCTTACTACTGCTGCTTTCATTTTCTCCTCTTTTTATGATTTATTTTGTTCAGCAAATAGTTTGC-3‘ |
| Alcohol dehydrogenase (adhA) of Lactococcus lactis | |
| fw_kivD_adhA (69 bp) | 5‘-GCAAACTATTTGCTGAACAAAATAAATCATAAAAAGAGGAGAAAATGAAAGCAGCAGTAGTAAGACAC-3‘ |
| rev_pSB1C3_adhA (80 bp) | 5‘-GTGACACCTTGCCCTTTTTTGCCGGACTGCAGCGGCCGCTACTAGTATTATTTAGTAAAATCAATGACCATTCGGCCTTC-3‘ |
| fw_pSB1C3_adhA (66 bp) | 5‘-GATTTCTGGAATTCGCGGCCGCTTCTAGAGAAAGAGGAGAAAATGAAAGCAGCAGTAGTAAGACAC-3‘ |
| rev_adhA_pSB1C3 (56 bp) | 5‘-GTCTTACTACTGCTGCTTTCATTTTCTCCTCTTTCTCTAGAAGCGGCCGCGAATTC-3‘ |
| fw-pSB1C3-BBa_B0034-adhaA (66 bp) | 5‘-GATTTCTGGAATTCGCGGCCGCTTCTAGAGAAAGAGGAGAAAATGAAAGCAGCAGTAGTAAGACAC-3‘ |
| rev-adhA-BBa_B0034-pSB1C3-adhaA (56 bp) | 5‘-GTCTTACTACTGCTGCTTTCATTTTCTCCTCTTTCTCTAGAAGCGGCCGCGAATTC-3‘ |
| Alcohol dehydrogenase (adhA) of E. coli | |
| fw-pSB1C3-Ec_adh (59 bp) | 5‘-CTGGAATTCGCGGCCGCTTCTAGAGAAAGAGGAGAAAATGCAGAACATCATCCGAAAAG-3‘ |
| rv-pSB1C3-Ec_adh (48 bp) | 5‘-GACTGCAGCGGCCGCTACTAGTATTAGTGACGGAAATCAATCACCATG-3‘ |
| fw-adhA-pSBC1 (48 bp) | 5‘-CATGGTGATTGATTTCCGTCACTAATACTAGTAGCGGCCGCTGCAGTC-3‘ |
| rv-adhA-pSBC1 (51 bp) | 5‘-CGGATGATGTTCTGCATTTTCTCCTCTTTCTCTAGAAGCGGCCGCGAATTC-3‘ |
| Biosafety and antibiotic-free selection | |
| Deletion of D-alanin racemases (alr and dadX) | |
| alr_fwd_redET (72 bp) | 5‘-AATTAGGTAATTAAAGCAAACACTTATCAAGGAACACAAATGCAAGCGGCAATTAACCCTCACTAAAGGGCG-3‘ |
| alr_rev_redEt (73 bp) | 5‘-TTTTATTGGCCGGATAAGGCATTCACGCCGCATCCGGCACAGACAATCAATAATACGACTCACTATAGGGCTC-3‘ |
| dadX_fwd_redET (72 bp) | 5‘-CGTCCGGGCCATTTACATGGCGCACACAGCTAAGGAAACGAGATGACCCAAATTAACCCTCACTAAAGGGCG-3‘ |
| dadX_rev_redET (73 bp) | 5‘-TGATTTTTTTGCACCCAGAAGACGTTGCCTCCGATCCGGCTTACAACAAGTAATACGACTCACTATAGGGCTC-3‘ |
| alr_Ec_control1 (20 bp) | 5‘-GCTGGAGATGCCATCAGAAC-3‘ |
| alr_Ec_control2 (20 bp) | 5‘-CCGGCGAATATTGCTACGTG-3‘ |
| dadX_Ec_control1 (20 bp) | 5‘-GCTTTAATACGCCCGTTGAC-3‘ |
| dadX_Ec_control2 (20 bp) | 5‘-CTGGATCAACGCTTCTTTGG-3‘ |
| Replacement of the canamycin resistance (CM) by the alr | |
| pSB1C3_Cm_del_fwd (40 bp) | 5‘-ACAACAGTTGCCGCTTGCATTTTAGCTTCCTTAGCTCCTG-3‘ |
| pSB1C3_Cm_del_rev (41 bp) | 5‘-CGATGAAATACGTGGATTAATTTGATATCGAGCTCGCTTGG-3‘ |
| alr_Cm_del_fwd (31 bp) | 5‘-GGAAGCTAAAATGCAAGCGGCAACTGTTGTG-3‘ |
| alr_Cm_del_rev (31 bp) | 5‘-CGATATCAAATTAATCCACGTATTTCATCGC-3‘ |
| Konstitutive expression of alr for plasmid copy number for the antibiotic-free selection | |
| pSB_kon_pre_GFP (42 bp) | 5‘-AGTTCTTCTCCTTTACGCATTTTCTCCTCTTTCTCTAGTAGC-3‘ |
| pSB_kon_suf_GFP (43 bp) | 5‘-ACTACGCTTTAGTAGCTTAATACTAGTAGCGGCCGCTGCAGTC-3‘ |
| AB_kon_GFP_fwd (32 bp) | 5‘-AAAGAGGAGAAAATGCGTAAAGGAGAAGAACT-3‘ |
| AB_kon_GFP_rev (34 bp) | 5‘-GCTACTAGTATTAAGCTACTAAAGCGTAGTTTTC-3‘ |
| LVA-tag for the D-alanin racemase | |
| alr_LVA_fwd (54 bp) | 5‘-GCTACTAAAGCGTAGTTTTCGTCGTTTGCAGCGGTCGCGATGAAATACGTGGAT-3‘ |
| alr_LVA_rev (52 bp) | 5‘-AACGACGAAAACTACGCTTTAGTAGCTTAATATTTGATGCCTGGCTCTAGTA-3‘ |
