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Team:Saarland/Test
From 2014.igem.org
Materials
1. Chemicals
| Chemical | Manufacturer |
|---|---|
| 2 log DNA ladder | New England Biolabs |
| 2-Mercapto Ethanol | Merck |
| Acetic Acid 100% | zChL UdS |
| Agar Kobe I | Roth |
| Agarose | Peqlab, Biozym |
| Ampicillin | Roth |
| APS (Ammoniumpersulfat) | zChL UdS |
| BactoTM Casamino-acids | BD |
| BactoTM Tryptone | BD |
| BactoTM Yeast-Extract | BD |
| Boric Acid | Merck |
| Bovine Serum Albumin (BSA) | New England Biolabs |
| Bromophenol Blue | Merck |
| Calcium Chloride | Calbiochem |
| Chloramphenicol | Fagran |
| D(+)-Glucose | Sigma Aldrich |
| ddH2O (MilliQ) (MILLIPORE 0,22 μm Membranfilter) | Biocel |
| DifcoTM Antibiotic Medium No. 3 | BD |
| Dimethylsulfoxide (DMSO) | Sigma Aldrich |
| dNTP-Mix (10 mM) | Peqlab |
| EDTA | Roth |
| Ethanol, HPLC grade | Sigma Aldrich |
| Glutamate | Sigma Aldrich |
| Glycerol | zChL UdS |
| Hydrochloric Acid 37 % (HCl) | VWR |
| Immersion Oil | Leica |
| Isopropanol | Sigma Aldrich |
| LB-Medium (Miller) | Sigma Aldrich |
| Magnesium Chloride Hexahydrate | VWR |
| Magnesium Sulfate | zChL UdS |
| Maleic Acid | Roth |
| Natriumchlorid | VWR |
| Natriumhydroxid | Grüssing |
| PEG-6000 | Merck |
| peqGREEN | Peqlab |
| Potassium Chloride | Grüssing |
| Potassium Dihydrogen Orthophosphate | Grüssing |
| Potassium Hydrogen Phosphate | Grüssing |
| Potassium Sulfate | zChL UdS |
| Proline | Merck |
| Protein Standard | New England Biolabs |
| Rotiphorese Gel 30 | Roth |
| SDS | Applichem/Roth |
| Succrose | Sigma Aldrich |
| Temed | Applichem |
| Tetracycline | Serva |
| Tris-Baze | Sigma Aldrich |
| Trizma Base | Sigma Aldrich |
| Xylose | Sigma Aldrich |
2. Enzymes
| Enzyme | Manufacturer |
|---|---|
| Lysozyme | Serva |
| T4 Ligase | Thermo Scientific |
| Taq-Polymerase | New England Biolabs |
| Q5 High Fidelity Polymerase (Master Mix) | New England Biolabs |
| FastDigest MluI | Thermo Scientific |
| FastDigest SacI | Thermo Scientific |
| FastDigest SphI | Thermo Scientific |
| FastDigest EcoRI | Thermo Scientific |
| FastDigest XbaI | Thermo Scientific |
| FastDigest SpeI | Thermo Scientific |
| FastDigest PstI | Thermo Scientific |
| Fast AP | Thermo Scientific |
3. oligonucleotides
| Abbreviation | 5´-Sequence-3´ |
|---|---|
| Generation of has2-gfp fusion gene | |
| SOE Has for: | TGTACACGCGTAAATCAAGGAGGTG |
| SOE Has rev: | CTCGCCCTTGCTCACCATTACATCTAATACCATATCGTATTGTTGACC |
| SOE GFP for: | ATGGTATTAGATGTAATGGTGAGCAAGGGCGAG |
| SOE GFP rev: | TGTACGAGCTCTTACTTGTACAGCTCGTCCATGC |
| Amplification of genomic DNA of B. megaterium | |
| RBS UDP-GlcDH for | TGTACGAGCTCAAATCAAGGAGGTGAATATACAATGACTAATATCACAGTAGCGGGTACTGG |
| UDP-GlcDH rev: | TGTACGCATGCTTAGACATTAGCTTTAATTTTAGCTTTGTATCC |
| Glk for | TGTAC GAATTC ATGGATGACAAATGGTTAG |
| Glk rev: | TGTAC CTGCAG TTA TTATTCAAAATAAGAT |
| PgmB for | TGTAC GAATTC ATGAAAAAAGAAGAGCTAC |
| PgmB rev: | TGTAC ACTAGT TTA TTAATTTTTTGTATAC |
| GtaB for | TGTAC GAATTC ATGACGATAAAAAAGGCAG |
| GtaB rev: | TGTAC CTGCAG TTA TTAGCTGAAGTTGTTTGC |
| Pgi for | TGTAC TCTAGA ATGACACATGTTCGTTTTG |
| Pgi rev: | TGTAC CTGCAG TTA TTATTTCAAACGCTTTTC |
| GlmS for | TGTAC TCTAGA ATGTGCGGAATTGTAGG |
| GlmS rev: | TGTAC CTGCAG TTA TTATTCTACAGTAACTG |
| GlmM for | TGTAC GAATTC ATGGGTAAGTATTTTGG |
| GlmM rev: | TGTAC CTGCAG TTA TTACTCTAAGCCCATTTC |
| GcaD for | TGTAC GAATTC ATGTCAAAAAGATATGC |
| GcaD rev: | TGTAC CTGCAG TTA TTAGGATTTTTTATTAATATC |
| Quick Change PCR for restriction site mutagenesis | |
| QC Has for: | GAATTTGTAGAAGATTGGTACTCTCAAGAATTTATGGGTAACCAG |
| QC has rev: | AAATTCTTGAGAGTACCAATCTTCTACAAATTCGTGTAATAAAGAG |
| QC UDP-GlcDH for: | TAATTATTAAAAACTCTGATGCTGGTAAATTATTTTTCACAACTG |
| QC UDP-GlcDH rev: | ACCAGCATCAGAGTTTTTAATAATTAATTCATCAAGCCCAGG |
| QC PgmB for: | TTGCAGCTGCTGATTGAGTTCTTCAG |
| QC PgmB rev: | CTGCAAGGGATCAGCCGAGTG |
| QC Pgi for: | AAATTCTTCTTTGTCATACTCCGTAGG |
| QC Pgi rev: | GAATTTGCACGCATTCAAAAAAGTGC |
| QC GlmS for: | AAATTCACTTGCTACGTGCACTTCTAC |
| QC GlmS rev: | GAATTTTCTTATAACATGCCGCTTTTATCTG |
| QC GlmM for: | TCGAGATCTGCAAATAAATGTGCTGC |
| QC GlmM for: | TCTCGATGCAGATATCTCTACAATGG |
| Sequencing | |
| pSB1C3 check for: | TGCCACCTGACGTCTAAGAAACC |
| pSB1C3 check rev: | ATTACCGCCTTTGAGTGAGCTG |
4. Media and Solutions
| Solution | Ingredients | Quantity |
|---|---|---|
| 10 x KPP Buffer | KH₂PO₄ K₂HPO₄ | 2,71 g 12.5 g |
