IGEM:MIT/2006/Notebook/2006-7-18: Difference between revisions
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==osmY Primer Design Part 2== | ==osmY Primer Design Part 2== | ||
Truncated Forward Primer: 5'- GTT TCT TCG AAT TCG CGG CCG CTT CTA | Truncated Forward Primer: 5'- GTT TCT TCG AAT TCG CGG CCG CTT CTA G'''GCT TAT GTT TTC GCT GAT ATC CC''' - 3' | ||
Total Length: 51 bp | Total Length: 51 bp | ||
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hairpin deltaG: -2.66 kcal/mol | hairpin deltaG: -2.66 kcal/mol | ||
self dimer deltaG: - | self dimer deltaG: -22.78 kcal/mol | ||
Full Forward Primer: 5'- GTT TCT TCG AAT TCG CGG CCG CTT CTA | Full Forward Primer: 5'- GTT TCT TCG AAT TCG CGG CCG CTT CTA G'''CTG GCA CAG GAA CGT TAT C''' - 3' | ||
Total Length: 47 bp | Total Length: 47 bp | ||
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self dimer deltaG: -98.3 kcal/mol | self dimer deltaG: -98.3 kcal/mol | ||
Reverse Primer: 5'-GTT TCT TCC TGC AGC GGC CGC TAC TAG | Reverse Primer: 5'-GTT TCT TCC TGC AGC GGC CGC TAC TAG TA'''T TGT TAA ATA TAG ATC ACA ATT TTG AAA CCG'''- 3' | ||
Total Length: | Total Length: 60 bp | ||
Annealing Length: | Annealing Length: 31 bp | ||
GC Content: | GC Content: 25.8% | ||
Melting Temperature: | Melting Temperature: 53.5 degrees C | ||
hairpin deltaG: -2.86 kcal/mol | hairpin deltaG: -2.86 kcal/mol | ||
self dimer deltaG: - | self dimer deltaG: -25.98 kcal/mol | ||
heterodimer deltaG with Forward Primer: -99.44 kcal/mol | heterodimer deltaG with Forward Primer: -99.44 kcal/mol | ||
Revision as of 13:49, 18 July 2006
Transformants
- IK control did not grow, though BSMT and BAMT grew
- ATF1 and Top10 control grew as well
- Next step is liquid culture all of them, and miniprep on ATF1
- SAMTC connected to both RBSs grew along with SAMTA connected to both RBSs and the control.
osmY Primer Design Part 2
Truncated Forward Primer: 5'- GTT TCT TCG AAT TCG CGG CCG CTT CTA GGCT TAT GTT TTC GCT GAT ATC CC - 3'
Total Length: 51 bp
Annealing Length: 23 bp
GC Content: 43.5%
Melting Temperature: 54.0 degrees C
hairpin deltaG: -2.66 kcal/mol
self dimer deltaG: -22.78 kcal/mol
Full Forward Primer: 5'- GTT TCT TCG AAT TCG CGG CCG CTT CTA GCTG GCA CAG GAA CGT TAT C - 3'
Total Length: 47 bp
Annealing Length: 19 bp
GC Content: 52.6%
Melting Temperature: 53.4 degrees C
hairpin deltaG: -3.99 kcal/mol
self dimer deltaG: -98.3 kcal/mol
Reverse Primer: 5'-GTT TCT TCC TGC AGC GGC CGC TAC TAG TAT TGT TAA ATA TAG ATC ACA ATT TTG AAA CCG- 3'
Total Length: 60 bp
Annealing Length: 31 bp
GC Content: 25.8%
Melting Temperature: 53.5 degrees C
hairpin deltaG: -2.86 kcal/mol
self dimer deltaG: -25.98 kcal/mol
heterodimer deltaG with Forward Primer: -99.44 kcal/mol
heterodimer deltaG with Conserved Forward Primer: -99.97 kcal/mol
Meeting
- Tomorrow: Sequence: 4 SAMT-C connected to rbs & term-30, and 4 SAMT-C connected to rbs & term-32
- end of the day: Liquid culture all transformants (Duda. BAMT in indole-knockout, Duda. BSMT in indole-knockout, the two above)
- Make liquid culture from the glycerols of the constitutive promoters
- Veena: miniprep pCHBA, sequence it with primers from Reshma
- Andre- request plasmid for vanilla
- maybe: re-cut miniprepped BSMT
- research exponential phase promoter
Additional Smells
- Myrcene and Linalool have floral scents
- Myrcene synthase: http://content.febsjournal.org/cgi/content/full/268/21/5633
Annealing Parts for Synthesis of the osmY Promoter
Forward Strand:
5'-ctagaggcttatgttttcgctgatatcccgagcggtttcaaaattgtgatctatatttaacaaatactagtagcggccgctgca-3'
Length: 84 bases
GC Content: 41.7%
Melting Temperature: 69.7 degrees Celsius
Complementary Strand:
5'-GCGGCCGCTACTAGTATTTGTTAAATATAGATCACAATTTTGAAACCGCTCGGGATATCAGCGAAAACATAAGCCT-3'
Length: 76 bases
GC Content: 40.8%
Melting Temperature: 69.1 degrees Celsius
Heterodimer DeltaG: -161.93 kcal/mol
Also, I contacted Professor Schellhorn last night inquiring about a plasmid containing osmY or the sequences of the primers he used to PCR osmY out of E. coli.
