Template:SBB10 ConstructionFiles Chel sbb02: Difference between revisions
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==JCA Notes== | ==JCA Notes== | ||
*Have gtcc instead of atg in prefix location | *Have gtcc instead of atg in prefix location | ||
*Need to put part on Clotho | |||
*Otherwise correct | *Otherwise correct | ||
*Internal oligos are longer than necessary, but would work | *Internal oligos are longer than necessary, but would work | ||
*Due to similarity to other part, some oligos are being replaced: | *Due to similarity to other part, some oligos are being replaced: | ||
<pre> | <pre> | ||
Construction of | Construction of sbb02: N15 Protelomerase | ||
PCR MNF11-F/BDBn15001R on Bca1455 ( | PCR MNF11-F/CA1472 on Bca1455 1-3 (569bp, gp=A) | ||
PCR BDBn15002F/BDBn15002R on Bca1485 | PCR ca1461/BDBn15001R on Bca1455 1-1 (762bp, gp=B) | ||
PCR BDBn15002F/BDBn15002R on Bca1485 2-1 (1023 bp, gp=C) | |||
-------------------------------------------------------------- | -------------------------------------------------------------- | ||
PCR BDBn15001F/BDBn15002R on A+B ( | PCR BDBn15001F/BDBn15002R on A+B+C (1924 bp, EcoRI/BamHI) | ||
Sub in pBjk2741-Bca1144 (EcoRI/BamHI, 910+2170, L) | Sub in pBjk2741-Bca1144 (EcoRI/BamHI, 910+2170, L) | ||
Product is pBjk2741-sbb01 {rbs.part!} | Product is pBjk2741-sbb01 {rbs.part!} | ||
| Line 32: | Line 34: | ||
MNF11-F Forward PCR of Part 1 of sbb02 | MNF11-F Forward PCR of Part 1 of sbb02 | ||
ccagtGAATTCatgAGATCTAGCAAGGTAAAAATCGGTGAGTTG | ccagtGAATTCatgAGATCTAGCAAGGTAAAAATCGGTGAGTTG | ||
CA1461 GGAAAGGATTGCAGAAAAGAATAACCGCGAATACTTTTAACGCCTATATGA | |||
PCA assembly of protelomerasefiveprime (Bca1455) | |||
CA1472 TTGTTGGAATAACTTATAAAGATAGTCACGGCGCTCCTTCCAATCATCACT | |||
PCA assembly of protelomerasefiveprime (Bca1455) | |||
BDBn15001R Reverse retrieval and SOE-site introduciton on Bca1455 | BDBn15001R Reverse retrieval and SOE-site introduciton on Bca1455 | ||
CGCTTTTATCTTCACTGCGTTTTTTAGCTTGCCCTGAG | CGCTTTTATCTTCACTGCGTTTTTTAGCTTGCCCTGAG | ||
| Line 37: | Line 43: | ||
CTCAGGGCAAGCTAAAAAACGCAGTGAAGATAAAAGCG | CTCAGGGCAAGCTAAAAAACGCAGTGAAGATAAAAGCG | ||
BDBn15002R Reverse retrieval and BioBricking on Bca1485 | BDBn15002R Reverse retrieval and BioBricking on Bca1485 | ||
CTGATggatccttaGCTGTAGTACGTTTCCCATGCGG | |||
part: | part: | ||
GATCTAGCAAGGTAAAAATCGGTGAGTTGATCAACACGCTTGTGAATGAGGTAGAGGCCATTGATGCCTCAGACCGCCCACAAGGCGACAAAACGAAGAGAATTAAAGCCGCAGCCGCACGGTATAAGAACGCGTTATTTAATGATAAAAGAAAGTTCCGTGGGAAAGGATTGCAGAAAAGAATAACCGCGAATACTTTTAACGCCTATATGAGCAGGGCAAGAAAGCGGTTTGATGATAAATTACATCATAGCTTTGATAAAAATATTAATAAATTATCGGAAAAGTATCCTCTTTACAGCGAAGAATTATCTTCATGGCTTTCTATGCCTACGGCTAATATTCGCCAGCACATGTCATCGTTACAATCTAAATTGAAAGAAATAATGCCGCTTGCCGAAGAGTTATCAAATGTAAGAATAGGCTCTAAAGGCAGTGATGCAAAAATAGCAAGACTAATAAAAAAATATCCAGATTGGAGTTTTGCTCTTAGTGATTTAAACAGTGATGATTGGAAGGAGCGCCGTGACTATCTTTATAAGTTATTCCAACAAGGCTCTGCGTTGTTAGAAGAACTACACCAGCTCAAGGTCAACCATGAGGTTCTGTACCATCTCCAGCTAAGCCCTGCGGAGCGTACATCTATACAGCAACGATGGGCCGATGTTCTGCGCGAGAAGAAGCGTAATGTTGTGGTTATTGACTACCCAACATACATGCAGTCTATCTATGATATTTTGAATAATCCTGCGACTTTATTTAGTTTAAACACTCGTTCTGGAATGGCACCTTTGGCCTTTGCTCTGGCTGCGGTATCAGGGCGAAGAATGATTGAGATAATGTTTCAGGGTGAATTTGCCGTTTCAGGAAAGTATACGGTTAATTTCTCAGGGCAAGCTAAAAAACGCAGTGAAGATAAAAGCGTAACCAGAACGATTTATACTTTATGCGAAGCAAAATTATTCGTTGAATTATTAACAGAATTGCGTTCTTGCTCTGCTGCATCTGATTTCGATGAGGTTGTTAAAGGATATGGAAAGGATGATACAAGGTCTGAGAACGGCAGGATAAATGCTATTTTAGCAAAAGCATTTAACCCTTGGGTTAAATCATTTTTCGGCGATGACCGTCGTGTTTATAAAGATAGCCGCGCTATTTACGCTCGCATCGCTTATGAGATGTTCTTCCGCGTCGATCCACGGTGGAAAAACGTCGACGAGGATGTGTTCTTCATGGAGATTCTCGGACACGACGATGAGAACACCCAGCTGCACTATAAGCAGTTCAAGCTGGCCAACTTTTCCAGAACCTGGCGACCGGAAGTTGGGGATGAAAACACCAGGCTGGTGGCTCTCCAGAAACTGGACGATGAAATGCCAGGCTTTGCCAGAGGTGACGCTGGCGTCCGTCTCCATGAAACCGTTAAGCAGCTGGTGGAGCAGGACCCATCAGCAAAAATAACCAACAGCACTCTCCGGGCCTTTAAATTTAGCCCGACGATGATTAGCCGGTACCTGGAGTTTGCCGCTGATGCATTGGGGCAGTTCGTTGGCGAGAACGGGCAGTGGCAGCTCAAGATAGAGACACCTGCAATCGTCCTGCCTGATGAAGAATCCGTTGAGACCATCGACGAACCGGATGATGAGTCCCAAGACGACGAGCTGGATGAAGATGAAATTGAGCTCGACGAGGGTGGCGGCGATGAACCAACCGAAGAGGAAGGGCCAGAAGAACATCAGCCAACTGCTCTAAAACCCGTCTTCAAGCCTGCAAAAAATAACGGGGACGGAACGTACAAGATAGAGTTTGAATACGATGGAAAGCATTATGCCTGGTCCGGCCCCGCCGATAGCCCTATGGCCGCAATGCGATCCGCATGGGAAACGTACTACAGCTAAG | |||
</pre> | </pre> | ||
</blockquote> | </blockquote> | ||
__NOTOC__ | __NOTOC__ | ||
Latest revision as of 16:48, 22 February 2010
sbb02: N15 Protelomerase
PCR MNF11-F/MNF12-R on pBca9523-Bca1455 (945 bp, gp = A)
PCR MNF13-F/MNF14-R on pBca9523-Bca1485 (1033 bp, gp = B)
-------------
PCR MNF11-F/MNF14-R on A + B (1925 bp, EcoRI/BamHI)
Sub in pBjk2741-Bca1144 (EcoRI/BamHI, 910+2170, L)
Product is pBjk2741-sbb02 {<protel!}
-------------
MNF11-F Forward PCR of Part 1 of sbb02 ccagtGAATTCgtccAGATCTAGCAAGGTAAAAATCGGTGAGTTG
MNF12-R SOEing of Part1 of sbb02 ggttacgcttttatcttcactgcgTTTTTTAGCTTGCCCTGAGAAATTAACCG
MNF13-F SOEing of Part2 of sbb02 CGGTTAATTTCTCAGGGCAAGCTAAAAAAcgcagtgaagataaaagcgtaacc
MNF14-R Reverse PCR of Part 2 of sbb02 gcagtggatccTCAGCTGTAGTACGTTTCCCATGC
JCA Notes
- Have gtcc instead of atg in prefix location
- Need to put part on Clotho
- Otherwise correct
- Internal oligos are longer than necessary, but would work
- Due to similarity to other part, some oligos are being replaced:
Construction of sbb02: N15 Protelomerase PCR MNF11-F/CA1472 on Bca1455 1-3 (569bp, gp=A) PCR ca1461/BDBn15001R on Bca1455 1-1 (762bp, gp=B) PCR BDBn15002F/BDBn15002R on Bca1485 2-1 (1023 bp, gp=C) -------------------------------------------------------------- PCR BDBn15001F/BDBn15002R on A+B+C (1924 bp, EcoRI/BamHI) Sub in pBjk2741-Bca1144 (EcoRI/BamHI, 910+2170, L) Product is pBjk2741-sbb01 {rbs.part!} MNF11-F Forward PCR of Part 1 of sbb02 ccagtGAATTCatgAGATCTAGCAAGGTAAAAATCGGTGAGTTG CA1461 GGAAAGGATTGCAGAAAAGAATAACCGCGAATACTTTTAACGCCTATATGA PCA assembly of protelomerasefiveprime (Bca1455) CA1472 TTGTTGGAATAACTTATAAAGATAGTCACGGCGCTCCTTCCAATCATCACT PCA assembly of protelomerasefiveprime (Bca1455) BDBn15001R Reverse retrieval and SOE-site introduciton on Bca1455 CGCTTTTATCTTCACTGCGTTTTTTAGCTTGCCCTGAG BDBn15002F Forward retrieval and SOE-site introduciton on Bca1485 CTCAGGGCAAGCTAAAAAACGCAGTGAAGATAAAAGCG BDBn15002R Reverse retrieval and BioBricking on Bca1485 CTGATggatccttaGCTGTAGTACGTTTCCCATGCGG part: GATCTAGCAAGGTAAAAATCGGTGAGTTGATCAACACGCTTGTGAATGAGGTAGAGGCCATTGATGCCTCAGACCGCCCACAAGGCGACAAAACGAAGAGAATTAAAGCCGCAGCCGCACGGTATAAGAACGCGTTATTTAATGATAAAAGAAAGTTCCGTGGGAAAGGATTGCAGAAAAGAATAACCGCGAATACTTTTAACGCCTATATGAGCAGGGCAAGAAAGCGGTTTGATGATAAATTACATCATAGCTTTGATAAAAATATTAATAAATTATCGGAAAAGTATCCTCTTTACAGCGAAGAATTATCTTCATGGCTTTCTATGCCTACGGCTAATATTCGCCAGCACATGTCATCGTTACAATCTAAATTGAAAGAAATAATGCCGCTTGCCGAAGAGTTATCAAATGTAAGAATAGGCTCTAAAGGCAGTGATGCAAAAATAGCAAGACTAATAAAAAAATATCCAGATTGGAGTTTTGCTCTTAGTGATTTAAACAGTGATGATTGGAAGGAGCGCCGTGACTATCTTTATAAGTTATTCCAACAAGGCTCTGCGTTGTTAGAAGAACTACACCAGCTCAAGGTCAACCATGAGGTTCTGTACCATCTCCAGCTAAGCCCTGCGGAGCGTACATCTATACAGCAACGATGGGCCGATGTTCTGCGCGAGAAGAAGCGTAATGTTGTGGTTATTGACTACCCAACATACATGCAGTCTATCTATGATATTTTGAATAATCCTGCGACTTTATTTAGTTTAAACACTCGTTCTGGAATGGCACCTTTGGCCTTTGCTCTGGCTGCGGTATCAGGGCGAAGAATGATTGAGATAATGTTTCAGGGTGAATTTGCCGTTTCAGGAAAGTATACGGTTAATTTCTCAGGGCAAGCTAAAAAACGCAGTGAAGATAAAAGCGTAACCAGAACGATTTATACTTTATGCGAAGCAAAATTATTCGTTGAATTATTAACAGAATTGCGTTCTTGCTCTGCTGCATCTGATTTCGATGAGGTTGTTAAAGGATATGGAAAGGATGATACAAGGTCTGAGAACGGCAGGATAAATGCTATTTTAGCAAAAGCATTTAACCCTTGGGTTAAATCATTTTTCGGCGATGACCGTCGTGTTTATAAAGATAGCCGCGCTATTTACGCTCGCATCGCTTATGAGATGTTCTTCCGCGTCGATCCACGGTGGAAAAACGTCGACGAGGATGTGTTCTTCATGGAGATTCTCGGACACGACGATGAGAACACCCAGCTGCACTATAAGCAGTTCAAGCTGGCCAACTTTTCCAGAACCTGGCGACCGGAAGTTGGGGATGAAAACACCAGGCTGGTGGCTCTCCAGAAACTGGACGATGAAATGCCAGGCTTTGCCAGAGGTGACGCTGGCGTCCGTCTCCATGAAACCGTTAAGCAGCTGGTGGAGCAGGACCCATCAGCAAAAATAACCAACAGCACTCTCCGGGCCTTTAAATTTAGCCCGACGATGATTAGCCGGTACCTGGAGTTTGCCGCTGATGCATTGGGGCAGTTCGTTGGCGAGAACGGGCAGTGGCAGCTCAAGATAGAGACACCTGCAATCGTCCTGCCTGATGAAGAATCCGTTGAGACCATCGACGAACCGGATGATGAGTCCCAAGACGACGAGCTGGATGAAGATGAAATTGAGCTCGACGAGGGTGGCGGCGATGAACCAACCGAAGAGGAAGGGCCAGAAGAACATCAGCCAACTGCTCTAAAACCCGTCTTCAAGCCTGCAAAAAATAACGGGGACGGAACGTACAAGATAGAGTTTGAATACGATGGAAAGCATTATGCCTGGTCCGGCCCCGCCGATAGCCCTATGGCCGCAATGCGATCCGCATGGGAAACGTACTACAGCTAAG
