Part:BBa_K4061040
eGFP antisense to downregulate eGFP (BBa_K4061002)
The mRNA antisense system has been known to downregulate gene expression at the translational level. The binding of antisense mRNA to the coding mRNA will cause degradation of the double-stranded mRNA. Thus, the coding mRNA is unable to get translated into protein, resulting in a declining protein expression. This particular part, eGFP antisense was made to downregulate the expression of eGFP (BBa_K4061002). We designed the eGFP antisense such that the antisense mRNA fragment will bind with the first 22 nucleotides sequence of the eGFP, the RBS (BBa_B0034), and 10 nucleotides sequence of the OmpF promoter (BBa_K116500). Therefore, the binding of the antisense mRNA will inhibit the ribosome from binding to the ribosome binding site.
Furthermore, our antisense eGFP construct was equipped with an Hfq binding site to stabilize the single strand mRNA fragment to protect it from degradation. The Hfq binding site that we utilize named Spot 42 has a high specificity to Hfq protein. Hfq protein is endogenous to E.coli and is known to interact with small antisense RNA that has AU rich region to modulate its stability.
In our context, the eGFP (BBa_K4061002) that we want to downregulate was controlled under an OmpF promoter (BBa_K116500). Thus, the respective mRNA strand would bind to a region of the OmpF promoter. For further utilization of this antisense fragment, it needs to be altered according to the context of your project.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 14
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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