Part:BBa_K1722011
shTERT+tRNA(CUA) Composite
hTERT, which is short for human telomerase reverse transcriptase, is a cancer-cell specific promoter. It can be activated inside cancer cells with no effect on normal cells. By mutating four base pairs of hTERT sequence, we achieved an improved promoter with higher promote efficiency named super hTERT(shTERT). As an important component of human telomerase, hTERT express only in tumor cells and other immortal cells which are telomerase positive. Only in tumor cells that can express TERT can the promoter being activated to realize targeted expression of effector gene.
We constructed another gene downstream of shTERT called tRNA, which encodes an unusual transfer RNA(tRNA) with a CUA anticodon. The tRNA(CUA) that is produced can be charged with Acetyllysine(Ack) with the help of Acetyllysine tRNA synthetase(AckRS). It is also the first step in translating UAG amber codon.
In our project, when shTERT is activated inside bladder cancer cell, downstream tRNA sequence can be expressed to produce tRNA with CUA as its anticodon. Together with AckRS, tRNA can carry Ack and pair with the amber UAG stop codon of the mRNA chain to completely produce the interest protein.
Sequence and Features
- 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
Both shTERT promoter and tRNA are achieved from Shenzhen Second People's Hospital. We designed primers and amplified the gene sequences from psi-Check2 vector. Using 3A Assembly method, we constructed hTERT and tRNA in pSB1C3.
Source
Both shTERT promoter and tRNA are achieved from Shenzhen Second People's Hospital.
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