Difference between revisions of "Part:BBa K1051800:Design"

 
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===Design Notes===
 
===Design Notes===
 
To mutation four Restriction Enzyme cutting sites(SpeI) and two codons(D10A & H841A),we design PCR primers to mutate them by PCR using Phution DNA polymerase.
 
To mutation four Restriction Enzyme cutting sites(SpeI) and two codons(D10A & H841A),we design PCR primers to mutate them by PCR using Phution DNA polymerase.
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Refrence http://www.ncbi.nlm.nih.gov/pubmed/?term=repurposing+CRISPR+as+an+RNA-Guided+Platfrom
 
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===Source===
 
===Source===
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===References===
 
===References===
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Repurposing CRISPR as an RNA-guided platform for sequence-specific control of gene expression [http://www.ncbi.nlm.nih.gov/pubmed/?term=repurposing+CRISPR+as+an+RNA-Guided+Platfrom]

Latest revision as of 17:01, 11 July 2013


protein dCas9.Two mutations,D10A and H841A,from Cas9 gene without stop codon.


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 2740
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 2886
    Illegal BsaI site found at 3740
    Illegal BsaI.rc site found at 1411


Design Notes

To mutation four Restriction Enzyme cutting sites(SpeI) and two codons(D10A & H841A),we design PCR primers to mutate them by PCR using Phution DNA polymerase.

Source

The Cas9 gene was got from Mr. Kai Tian,BGI Shenzhen China. The gene was splited into three parts to synthetize optimizing the codons in eucaryon.



References

Repurposing CRISPR as an RNA-guided platform for sequence-specific control of gene expression [http://www.ncbi.nlm.nih.gov/pubmed/?term=repurposing+CRISPR+as+an+RNA-Guided+Platfrom]