Part:BBa_K627007:Design
Fusion of c-myc-tag and gene III
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
Gene III was amplified from the vector pak100 bla KDIR using the following primer
Forward: TAAGCTTCTAGATGGCCGGCGAGCAGAAGCTGATCTCTGAGGAAGACCTGGGTGGTGGCTCTGGTTCC
Reverse: TGCTTAGACGTCCTGCAGCGGCCGCTACTAGTATTAACCGGTAGACTCCTTATTACGCAGTA
Because c-myc-gene III is a fusion part (RFC25), no start codon is needed. Before the c-myc sequence a NgoMIV restriction site is located which has compatible overhangs to AgeI restriction sites. The ligation of AgeI and NgoMIV overhangs will result in a scar coding for threonine and glycine. So genes of interest containing AgeI restriction sites can be easily fused to gene III without frame shift. The gene III sequence contains no stop codon because there is one located between AgeI and SpeI recognition site directly merged behind gene III sequence.
Source
The gene-III-protein is a coat protein from the filamentous bacteriophage M13.
References
Fuh G., Sidhu S.S. (2000). Efficient phage display of polypeptides fused to the carboxy-terminus of the M13 gene-3 minor coat protein. FEBS Lett. 480(2-3):231-4
Rakonjac J., Feng J., Model P. (1999). Filamentous phage are released from the bacterial membrane by a two-step mechanism involving a short C-terminal fragment of pIII. J Mol Biol. 289(5):1253-65
Smith, G.P. (1985). Filamentous fusion phage: Novel expression vectors that display cloned antigens on the virus surface. Science 228: 1315-17