Difference between revisions of "Part:BBa K1997010"

Latest revision as of 01:15, 21 October 2016

dCas9

This part codes dCas9 protein.

Usage and Biology

Functional elucidation of causal genetic variants and elements requires precise genome editing technologies. The type II prokaryotic CRISPR (clustered regularly interspaced short palindromic repeats)/Cas adaptive immune system has been shown to facilitate RNA-guided site-specific DNA cleavage. Cas9 can also be converted into a nicking enzyme to facilitate homology-directed repair with minimal mutagenic activity. Lastly, multiple guide sequences can be encoded into a single CRISPR array to enable simultaneous editing of several sites within the mammalian genome, demonstrating easy programmability and wide applicability of the RNA-guided nuclease technology. ¹

Sequence and Features

Sequence and Features

Experimental Validation

This part is validated through two ways: PCR and functional testing

PCR

Methods

The PCR is performed with Premix EX Taq by Takara.

F-Prime: 5’- GAATTCGCGGCCGCTTCTAGAATGC-3’

R-Prime: 5’- GGACTAGTATTATTGTTTGTCTGCC-3’

The PCR protocol is selected based on the Users Manuel. The Electrophoresis was performed on a 1％ Agarose glu. The result of the agarose electrophoresis was shown on the picture below.

Functional Test

This part is tested in the composite part BBa_K1997024 and BBa_K1997025.

References

[1] Cong, L.et al.Multiplex Genome Engineering Using CRISPR/Cas Systems,Science 339, 819-823,(2013)