Difference between revisions of "Part:BBa K4329004"

Latest revision as of 12:22, 10 October 2022

Knockdown of Bifunctional BirA enzyme

What it is?

By using the CRISPRi system, we are able to decrease the expression of the Bifunctional BirA enzyme which is an enzyme to consume the biotin in E. coli. In our project, we are aiming at biosynthesizing biotin through the metabolic pathways to generate more biotin in E. coli.

Usage and Biology

With the CRISPRi/dCas9 system targeting the bifunctional BirA enzyme after the production of biotin, the dCas9 system will attach to the enzyme, blocking other DNA from combining with the enzyme. Thus, the enzyme would be in a temporarily dysfunctional state.

Characterization

Fig.1 CRISPRi/dCas9 system targeting the bifunctional BirA enzyme can block down the expression of BirA and lead to generating more biotin in E. coli. (A)Primer designed for Bifunctional BirA enzyme in the region 200-300. (B)Electrophoresis of BirA enzyme with and without dCas9 system.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
INCOMPATIBLE WITH RFC[12]
Illegal NheI site found at 1202
Illegal NheI site found at 4217
Illegal NheI site found at 4240
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
INCOMPATIBLE WITH RFC[25]
Illegal AgeI site found at 4264
1000
INCOMPATIBLE WITH RFC[1000]
Illegal BsaI.rc site found at 49

@@ Line 9: / Line 9: @@
 ===Usage and Biology===
-With the CRISPRi/dCas9 system targeting the bifunctional BirA enzyme after the production of biotin, the dCas9 system will attach to the enzyme, blocking other DNA from combining with the enzyme. Thus, the enzyme would be in a temporary dysfunctional state.
+With the CRISPRi/dCas9 system targeting the bifunctional BirA enzyme after the production of biotin, the dCas9 system will attach to the enzyme, blocking other DNA from combining with the enzyme. Thus, the enzyme would be in a temporarily dysfunctional state.
 ===Characterization===
-[[File:004 result.png|700px|thumb|center|Fig.4  CRISPRi/dCas9 system targeting the bifunctional BirA enzyme can block down the expression of BirA and lead to generating more biotin in E. coli. (A)Primers designed for Bifunctional BirA enzyme in the region 200-300 and 400-600. (B)Electrophoresis of BirA enzyme with and without dCas9 system.]]
+[[File:004 result2.png|700px|thumb|center|Fig.1  CRISPRi/dCas9 system targeting the bifunctional BirA enzyme can block down the expression of BirA and lead to generating more biotin in E. coli. (A)Primer designed for Bifunctional BirA enzyme in the region 200-300. (B)Electrophoresis of BirA enzyme with and without dCas9 system.]]