NYMU Taipei/Protocols2/Signal peptides and insulin A, B chain

From 2007.igem.org

BC005255: http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?db=nuccore&id=13528923

AAG TCTAGA aaagaggagaaa ACTAG                                                              | 5' ext + xba1 +rbs+ spacer
atggacaaattcgacgctaatcgccgcaaattgctggcgcttggtggcgttgcactcggtgccgccatcctgccgacccctgcgtttgca | ycbK TAT signal peptide
GGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAGAACTACTGCAACTAGTAG                      | insulin A chain
tactagtagcggccgctgcag                                                                      | spe1+not1+pst1 site
GAA                                                                                        | 3' extending region 

AAG TCTAGA aaagaggagaaa ACTAG                                                                    | 5' ext+ xba1 +rbs+ spacer
atggacaaattcgacgctaatcgccgcaaattgctggcgcttggtggcgttgcactcggtgccgccatcctgccgacccctgcgtttgca       | ycbK TAT signal peptide
TTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCTAGTAG | insulin B chain
tactagtagcggccgctgcag                                                                            |spe1+not1+pst1 site
GAA                                                                                              | 3' extending region

Common primer:

1.       TAT_Forward: AAG TCTAGA aaagaggagaaa ACTAG atggacaaattcgacgct

For TAT-insulin A:

2.  InsA_TAT_Reverse: ggctggggacgcaaacgtCCGTAACACCTTGTTACG
   

3. InsA_InsA_Forward: ccgacccctgcgtttgcaGGCATTGTGGAACAATGC

4.      InsA_Reverse: TTGATGACGTTGATCATCatgatcatcgccggcgacgtcCTT
   

For TAT-insulin B:

5.  InsB_TAT_Reverse: ggctggggacgcaaacgtAAACACTTGGTTGTGGAC
   

6. InsB_InsB_Forward: ccgacccctgcgtttgcaTTTGTGAACCAACACCTG

7.      InsB_Reverse: TGTGGGTTCTGGATCATCatgatcatcgccggcgacgtcCTT
   

File:Rbs tat ins.jpg