IGEM:MIT/2008/Notebook/Yogurt/sequences

Primers

Include sequence, usage, temperatures, concentration, start date.

P1: Forward Primer for Promoter

• • foward sequence: CCG CTT CTA GAG TAA TAC GAC TCA CTA TAG GGA ATA CAA GCT ACT TGT TCT TTT TGC ATA CTA GAG ATT AAA GAG GAG AAA TAC TAG ATG CAG AAG AAA AAA TCC GCA CGA TGC AGA AGA AAA AAT CCG C
  • length = 110 bp
  • melting temp = 68.8 C
  • GC content % = 37.3

P2: Reverse Primer for Peptide

• • reverse sequence: G TTC TTC TCC TTT ACG CAT AG AGC CTC CAC CAG AGC CTC
  • length = 69 bp
  • melting temp = 71.8 C
  • GC content % = 50.7

TEV Protease cut site

• Amino Acid Sequence : Glu-Asn-Leu-Tyr-Phe-Gln-Gly (Cuts between Gln-Gly)
• Base Pair Sequence: GAAAACCTGTACTTCCAGGGT

P3: Forward Primer for GFP

• • forward sequence: GA GGC TCT GGT GGA GGC TCT ATG CGT AAA GGA GAA GAA C

• • includes 21 bp of 2nd half of linker, and 19 bp of GFP
  • length = 40 bp
  • melting temp = 66.2 C
  • GC content % = 50%

P4: Reverse Primer for GFP

• • reverse sequence: CTG CAG CGG CCG CTA CTA GTA AGA GAA TAT AAA AAG CCA GAT TAT TAA TCC GGC TTT TTT ATT ATT TTT ATT AGT GGT GAT GGT GAT GAT GTT TGT ATA GTT CAT CCA TGC
  • length = 111 bp
  • melting temp = 69.6 C
  • GC content % = 36.0

P5: Reverse Primer for GFP without Terminator

• • reverse sequence: CTG CAG CGG CCG CTA CTA GTA TTA TTA GTG GTG ATG GTG ATG ATG TTT GTA TAG TTC ATC CAT GC

Plasmids

Include sequence, BioBrick #, functional features, digestion map.

PCR Products

Include sequence, template and primers, functional features.

Other

DNA sequence to synthesize - epitope/p1025/linker (6/17/08)

• The signal peptide sequence can be found at the NCBI genome database

• 6/17, updated signal peptide, silent mutations in 1st part of linker (GGC instead of GGT coding for Gly)

sequence: ATGCAGAAGAAAAAATCCGCACGCCATTTGAACAAAGTGGCTGAA TTAGCCGCAGCACTGCTCCTATCAGCGAGTCCACTGGCGGGAACTTTC GACTACAAAGACGACGACGACAAAGGTGGTGGTCAGCTGAAAACCGCT GACCTGCCGGCTGGTCGTGACGAAACCACCTCTTTCGTTCTGGTTGGT GGTGGTTACCCGTACGACGTTCCGGACTACGCTGGCGGCGGCTCTGGC GGCGGCTCTGGCGGCGGCTCTGAAAACCTGTACTTCCAGGGTGGTGGT GGTTCTGGTGGTGGTTCTGGTGGTGGTTCT

Whole Sequence

CCGCTTCTAGAGTAATACGACTCACTATAGGGAATACAAGCTACTTGTTC TTTTTGCATACTACAGATTAAAGAGGAGAAATACTAGATGATGCAGAAGA AAAAATCCGCACGCCATTTGAACAAAGTGGCTGAATTAGCCGCAGCACTG CTCCTATCAGCGAGTCCACTGGCGGGAACTTTCCAGTCAGCCGCTTTTGT CCAAGCTGCCAGTCAAGAAACGGACTACAAAGACGACGACGACAAAGGTG GTGGTCAGCTGAAAACCGCTGACCTGCCGGCTGGTCGTGACGAAACCACC TCTTTCGTTCTGGTTGGTGGTGGTTACCCGTACGACGTTCCGGACTACGC TGGTGGTGGTTCTGGTGGTGGTTCTGGTGGTGGTTCTGAAAACCTGTACT TCCAGGGTGGTGGTGGTTCTGGTGGTGGTTCTGGTGGTGGTTCTATGCGT AAAGGAGAAGAACTTTTCACTGGAGTTGTCCCAATTCTTGTTGAATTAGA TGGTGATGTTAATGGGCACAAATTTTCTGTCAGTGGAGAGGGTGAAGGTG ATGCAACATACGGAAAACTTACCCTTAAATTTATTTGCACTACTGGAAAA CTACCTGTTCCATGGCCAACACTTGTCACTACTTTCGGTTATGGTGTTCA ATGCTTTGCGAGATACCCAGATCATATGAAACAGCATGACTTTTTCAAGA GTGCCATGCCCGAAGGTTATGTACAGGAAAGAACTATATTTTTCAAAGAT GACGGGAACTACAAGACACGTGCTGAAGTCAAGTTTGAAGGTGATACCCT TGTTAATAGAATCGAGTTAAAAGGTATTGATTTTAAAGAAGATGGAAACA TTCTTGGACACAAATTGGAATACAACTATAACTCACACAATGTATACATC ATGGCAGACAAACAAAAGAATGGAATCAAAGTTAACTTCAAAATTAGACA CAACATTGAAGATGGAAGCGTTCAACTAGCAGACCATTATCAACAAAATA CTCCAATTGGCGATGGCCCTGTCCTTTTACCAGACAACCATTACCTGTCC ACACAATCTGCCCTTTCGAAAGATCCCAACGAAAAGAGAGACCACATGGT CCTTCTTGAGTTTGTAACAGCTGCTGGGATTACACATGGCATGGATGAAC TATACAAACATCATCACCATCACCACTAATAAAAATAATAAAAAAGCCGG ATTAATAATCTGGCTTTTTATATTCTCTTACTAGTAGCGGCCGCTGCAG