Part:BBa_K3982001

Cas14a1 expression gene

This sequence encodes the Cas14a1 protein. It is a highly compact RNA guided nuclease used by Team IISER_Berhampur 2021 in Project CODE M.

Usage and Biology

Cas14a1 (also called Un1Cas12f1) is a miniature cas protein used in CRISPR/Cas technology identified from archaebacteria and bacteriophages. It is a RNA guided nuclease that 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).

To know more about how this part is used in project CODE M, visit our wikipage. Team IISER_Berhampur 2021

Check composite parts BBa_K3982006, BBa_K3982025, BBa_K3982026 to see how this part has been utilised in our project.

References

1) Harrington, L. B., Burstein, D., Chen, J. S., Paez-Espino, D., Ma, E., Witte, I. P., Cofsky, J. C., Kyrpides, N. C., Banfield, J. F., & Doudna, J. A. (2018). Programmed DNA destruction by miniature CRISPR-Cas14 enzymes. Science (New York, N.Y.), 362(6416), 839–842. https://doi.org/10.1126/science.aav4294

2) Takeda, S. N., Nakagawa, R., Okazaki, S., Hirano, H., Kobayashi, K., Kusakizako, T., Nishizawa, T., Yamashita, K., Nishimasu, H., & Nureki, O. (2021). Structure of the miniature type V-F CRISPR-Cas effector enzyme. Molecular cell, 81(3), 558–570.e3. https://doi.org/10.1016/j.molcel.2020.11.035

3) Karvelis, T., Bigelyte, G., Young, J. K., Hou, Z., Zedaveinyte, R., Budre, K., Paulraj, S., Djukanovic, V., Gasior, S., Silanskas, A., Venclovas, Č., & Siksnys, V. (2020). PAM recognition by miniature CRISPR-Cas12f nucleases triggers programmable double-stranded DNA target cleavage. Nucleic acids research, 48(9), 5016–5023. https://doi.org/10.1093/nar/gkaa208

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal EcoRI site found at 1357
  Illegal PstI site found at 147
  Illegal PstI site found at 313
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 1357
  Illegal PstI site found at 147
  Illegal PstI site found at 313
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal EcoRI site found at 1357
  Illegal BamHI site found at 951
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal EcoRI site found at 1357
  Illegal PstI site found at 147
  Illegal PstI site found at 313
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal EcoRI site found at 1357
  Illegal PstI site found at 147
  Illegal PstI site found at 313
  Illegal AgeI site found at 607
  
• 1000
  COMPATIBLE WITH RFC[1000]