Part:BBa_K1943022
TRPC5-7
Transient receptor potential channel 5 (TRPC5) is a mechanosensitive (MS) channel which takes charge of hearing in our project. Future teams could use original TRPC5 plasmid (BBa_K1943021) to sense sound or other kinds of mechanical forces. Furthermore, we randomly mutated the ankyrin region of TRPC5 channel by PCR. We expected the ankyrin region to be a crucial part for sense mechanical forces based on the structure of TRPC5.(Detailed methods are shown in our notebook.)
These mutated TRPC5 plasmids(BBa_K1943022, BBa_K1943023, BBa_K1943024, BBa_K1943025, BBa_K1943026) have all been sequenced. Because of limited time, we have not yet transfected them into cells to observe whether the channels are more sensitive to sound and mechanical force or not. Future teams are welcomed to use and finish testing them.
The mutated amino acid sites are as listed:
| Part | Original amino acids | Amino acid after mutation |
|---|---|---|
| BBa_K1943022 | Trs(240)+Ala(27) | Glu(240)+Thr(27) |
| BBa_K1943023 | Ala(27) | Thr(27) |
| BBa_K1943024 | Met(134)+Ala(27) | Ile(134)+Thr(27) |
| BBa_K1943025 | Glu(236)+Ala(27) | Lys(236)+Thr(27) |
| BBa_K1943026 | Gln(128)+Ala(27) | Arg(128)+Thr(27) |
TRPC5 sequence contains 1 EcoRⅠ RE site and 2 PstⅠ RE sites, which are unacceptable for BioBricks. Therefore, we did site-mutation by PCR for three times. Detailed information are shown in our notebook.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 930
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 167
Illegal BglII site found at 1520 - 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 2539
Illegal BsaI.rc site found at 1627
Illegal BsaI.rc site found at 2054
Illegal SapI site found at 1722
| None |