Part:BBa_K3017065:Design

Construct for testing CRISPRi sgRNA suppression on fluorescence protein production

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal PstI site found at 3147
  Illegal PstI site found at 4569
  Illegal PstI site found at 4773
  Illegal PstI site found at 4803
  Illegal PstI site found at 6015
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 7
  Illegal NheI site found at 30
  Illegal NheI site found at 2131
  Illegal NheI site found at 2303
  Illegal NheI site found at 2326
  Illegal PstI site found at 3147
  Illegal PstI site found at 4569
  Illegal PstI site found at 4773
  Illegal PstI site found at 4803
  Illegal PstI site found at 6015
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 2608
  Illegal BamHI site found at 2070
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal PstI site found at 3147
  Illegal PstI site found at 4569
  Illegal PstI site found at 4773
  Illegal PstI site found at 4803
  Illegal PstI site found at 6015
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal PstI site found at 3147
  Illegal PstI site found at 4569
  Illegal PstI site found at 4773
  Illegal PstI site found at 4803
  Illegal PstI site found at 6015
  Illegal NgoMIV site found at 3435
  Illegal NgoMIV site found at 4539
  Illegal NgoMIV site found at 4612
  Illegal NgoMIV site found at 5097
  Illegal NgoMIV site found at 6006
  Illegal AgeI site found at 1905
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 705
  Illegal SapI site found at 1887
  

Source

dCas9 gene comes from Streptococcus pyogenes

References

Y. J. Lee, A. Hoynes-Oconnor, M. C. Leong, and T. S. Moon, “Programmable control of bacterial gene expression with the combined CRISPR and antisense RNA system,” Nucleic Acids Research, vol. 44, no. 5, pp. 2462–2473, Feb. 2016.

C. Anders, O. Niewoehner, A. Duerst, and M. Jinek, “Structural basis of PAM-dependent target DNA recognition by the Cas9 endonuclease,” Nature, vol. 513, no. 7519, pp. 569–573, 2014.

S. H. Sternberg, S. Redding, M. Jinek, E. C. Greene, and J. A. Doudna, “DNA Interrogation by the CRISPR RNA-Guided Endonuclease Cas9,” Biophysical Journal, vol. 106, no. 2, 2014.

T. Karvelis, G. Gasiunas, A. Miksys, R. Barrangou, P. Horvath, and V. Siksnys, “crRNA and tracrRNA guide Cas9-mediated DNA interference inStreptococcus thermophilus,” RNA Biology, vol. 10, no. 5, pp. 841–851, 2013.

T. Møller, T. Franch, P. Højrup, D. R. Keene, H. P. Bächinger, R. G. Brennan, and P. Valentin-Hansen, “Hfq,” Molecular Cell, vol. 9, no. 1, pp. 23–30, 2002.

G. M. Cech, A. Szalewska-Pałasz, K. Kubiak, A. Malabirade, W. Grange, V. Arluison, and G. Węgrzyn, “The Escherichia Coli Hfq Protein: An Unattended DNA-Transactions Regulator,” Frontiers in Molecular Biosciences, vol. 3, 2016.