Difference between revisions of "Part:BBa K1898100"
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We designed primers synthesized by Tri-I Biotech to remove a stop codon and to move CH25H cDNA to iGEM BioBrick. PCR was set up and the gel picture is shown above. The expected size for CH25H PCR check is ~1kb, which is shown in lane 2 and lane 3. | We designed primers synthesized by Tri-I Biotech to remove a stop codon and to move CH25H cDNA to iGEM BioBrick. PCR was set up and the gel picture is shown above. The expected size for CH25H PCR check is ~1kb, which is shown in lane 2 and lane 3. | ||
| − | + | <img src="https://parts.igem.org/File:Ch25h_gel.png"> | |
==Sequencing== | ==Sequencing== | ||
CH25H was back inserted into BBa_K880005 and sent to sequencing. The sequencing result shows that the CRYAA sequencing result is correct (with no stop codon): | CH25H was back inserted into BBa_K880005 and sent to sequencing. The sequencing result shows that the CRYAA sequencing result is correct (with no stop codon): | ||
Revision as of 08:54, 18 October 2016
ch25h, cholesterol 25 hydroxylase
This DNA codes for ch25h, cholesterol 25-hydroxylase, that catalyzes the formation of 25 hydroxycholesterol from cholesterol.
Usage and Biology
25 hydroxycholesterol (25 HC) has been reported to have a range of effects on the immune system, including the suppression of production of IgA by B cells and inducing the migration of human peripheral blood monocytes [1]. 25 HC is also found to be able to reverse protein aggregation and restore solubility to the proteins [2]. In our project, we aim to deliver CH25H to the lens, where 25 HC will be produced and reverse aggregated lens protein. Aged-induced cataracts occur when lens proteins get oxidized and clump together. The increase of 25HC in the lens will help to treat cataracts.
[1] Diczfalusy, U. (2013). On the formation and possible biological role of 25-hydroxycholesterol. Biochimie, 95(3), 455-460. doi:10.1016/j.biochi.2012.06.016
[2] Makley, L. N., McMenimen, K. A., DeVree, B. T., Goldman, J. W., McGlasson, B. N., Rajagopal, P., Dunyak, B.M., McQuade, T.J., Thompson, A.D., Sunahara, R., Klevit, R.E., Andley, U.P., and Gestwicki, J.E. (2015). Pharmacological chaperone for α-crystallin partially restores transparency in cataract models. Science, 350(6261), 674-677.
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]
Gel Pictures
We designed primers synthesized by Tri-I Biotech to remove a stop codon and to move CH25H cDNA to iGEM BioBrick. PCR was set up and the gel picture is shown above. The expected size for CH25H PCR check is ~1kb, which is shown in lane 2 and lane 3.
<img src="https://parts.igem.org/File:Ch25h_gel.png">
Sequencing
CH25H was back inserted into BBa_K880005 and sent to sequencing. The sequencing result shows that the CRYAA sequencing result is correct (with no stop codon):