Difference between revisions of "Part:BBa K1898100"

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<partinfo>BBa_K1898100 short</partinfo>
 
<partinfo>BBa_K1898100 short</partinfo>
  
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This DNA codes for ch25h, cholesterol 25-hydroxylase, that catalyzes the formation of 25 hydroxycholesterol from cholesterol.  
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This DNA codes for CH25H, also known as cholesterol 25-hydroxylase, which catalyzes the formation of 25 hydroxycholesterol (25HC) from cholesterol.  
  
 
===Usage and Biology===
 
===Usage and Biology===
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==Gel Pictures==
 
==Gel Pictures==
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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.
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We designed primers to remove a stop codon and to move CH25H cDNA into iGEM BioBrick. PCR was set up and the gel picture is shown below. The expected size for CH25H PCR check is ~1.1kb. The bands boxed in red are the correct bands.
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https://static.igem.org/mediawiki/parts/0/0e/Ch25h_gel.png
  
 
==Sequencing==
 
==Sequencing==
  
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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):
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CH25H was sent to sequencing and the result shows that the CH25H sequencing result is correct (with no stop codon):
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https://static.igem.org/mediawiki/parts/7/7e/Ch25h_sequence.png
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The four cutting sites are highlighted in red and CH25H is highlighted in orange

Latest revision as of 02:59, 20 October 2016


ch25h, cholesterol 25 hydroxylase

This DNA codes for CH25H, also known as cholesterol 25-hydroxylase, which catalyzes the formation of 25 hydroxycholesterol (25HC) 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


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]



Gel Pictures

We designed primers to remove a stop codon and to move CH25H cDNA into iGEM BioBrick. PCR was set up and the gel picture is shown below. The expected size for CH25H PCR check is ~1.1kb. The bands boxed in red are the correct bands.

Ch25h_gel.png

Sequencing

CH25H was sent to sequencing and the result shows that the CH25H sequencing result is correct (with no stop codon):

Ch25h_sequence.png

The four cutting sites are highlighted in red and CH25H is highlighted in orange