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Part:BBa_K1983004:Design
gp45 phenylalanine mutant M15 with C-terminal 6XHis-Tag
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal XhoI site found at 688
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
Designed including NdeI restriction site before the start codon (ATG) for efficient cloning into expression vectors, XhoI site for cutting between the protein and 6XHis-Tag.
Note: Due to technical issues, the part's suffix in the backbone is changed by one FIRST letter (T to G). However, it does not remove or add any other restriction sites and does not change the function of the suffix. The purpose of this note is to alert false negative results during sequencing if the part is used in the future.
Original suffix: TACTAGTAGCGGCCGCTGCAG Part suffix: GACTAGTAGCGGCCGCTGCAG
Gp45M15 mutation list
L220F; L23F; Y172F; I107F; M22F; M187F; Y55F; M194F; L197F; W199F; L167F; Y194F; Y39F; Y216F; M184F;
Primers
Primers used for amplification of the fragment:
Uni-FW: GTAGAATTCGCGGCCGCTTCTAG
Uni-RV: GTAGACTGCAGCGGCCGCTACTAG
Primers used for colony PCR screening:
For pETDuet-1:
Up-A1: GGATCTCGACGCTCTCCCT
Down3: ACCCCTCAAGACCCGTTTAG
Source
This part is derived T4 phage sliding clamp protein gp45 and was synthesized by Integrate DNA Technologies.