Part:BBa_K3268004
Strong expression of sfGFP in E. coli
To make a composite module that can highly express any inserted protein-coding genes, we had designed and constructed this strongly expressed sfGFP in E. coli as a proof of our concept.
1. The sfGFP coding region was PCR amplified from BBa_K1321337 using high-fidelity enzyme KOD-Plus.
2. The strong promoter along with BBa_B0034 RBS and BBa_B0015 transcriptional terminator on pSB1C3 plasmid backbone were PCR amplified from BBa_J364007 using high-fidelity enzyme KOD-Plus.
3. In order to make the strong promoter (along with BBa_B0034 RBS and BBa_B0015 transcriptional terminator on pSB1C3 plasmid backbone) useful to highly express inserted protein-coding genes, we had designed two PCR primers with HindII and BamHI sites respectively to generate a composite expression module without the original GFP coding sequence.
4. Any inserted protein-coding genes not containing HindII and BamHI sites in their coding sequences can then be amplified and cloned into our composite expression module improved from BBa_J364007.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 726
Illegal NheI site found at 749 - 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 1424
Illegal SapI.rc site found at 10
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