RNA

Part:BBa_K3982005

Designed by: Akankshya Ramkrishna Sahu   Group: iGEM21_IISER_Berhampur   (2021-10-01)
Revision as of 12:19, 11 October 2021 by Akankshya16 (Talk | contribs)


CODE M sgRNA for targeting mutant katG gene in Mycobacterium tuberculosis

Usage and Biology

Cas14a1 (also called as Un1Cas12f1) is a miniature cas protein used in CRISPR/Cas technology identified from archaebacteria and bacteriophages. It is a RNA guided nuclease which cleaves ssDNA without any Protospacer Adjacent Motif (PAM) specificity i.e. PAM independent ssDNA cleavage. This enables high-fidelity Single-nucleotide polymorphism (SNP) genotyping. It is guided by a single guide RNA (sgRNA) which has two components - CRISPR RNA (crRNA) and Trans-activating crispr RNA (tracrRNA).

crRNA is a 20bp nucleotide sequence complementary to the target DNA. tracrRNA is linked to the crRNA with a tetraloop that is responsible for cas14 activity. It plays a role in maturation of crRNA and has a stem-loop structure in itself that binds to the ribonuclease. Click here to know more about how this part is used in project CODE M by Team IISER_Berhampur 2021.

This sgRNA has been designed specifically for targeting the mutant katG gene along with Cas14a1 protein in Mycobacterium tuberculosis (Mtb) which confers to isoniazid drug resistance in Multidrug-resistant Tuberculosis (MDR-TB). The mutation appears at base number 11 (agc → acc) of the spacer (guiding) sequence. This is the S315T mutation in Mtb genome. (refer Ref 3) Refer to the wildtype katG targeting sgRNA here - BBa_K3982004

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]


[edit]
Categories
Parameters
None