Plasmid

Part:BBa_K4761429

Designed by: Pinhan Pan   Group: iGEM23_HiZJU-China   (2023-09-30)


pCas

pCas plasmid in CRISPR-Cas9 dual plasmid system. Expresses Cas9 endonuclease. It needs to be used in combination with the pTarget plasmid and Donor DNA to perform the gene knockout function. The vector has temperature-sensitive replication (RepA101ts), so when the pCas plasmid is converted into the bacteria, the bacteria must be grown at 30℃. Kanamycin resistance gene is carried on pCas plasmid. When we transform pCas plasmid into the receptive state DH5α, Kanamycin resistance gene can help us screen the transformed strains successfully.

Fig.1 Using Cas9 plasmid, paired with sgRNA expression box and Donor DNA to obtain ΔpheA, ΔpykA, ΔpykF E. coli strain mutants by CRISPR-Cas9 gene knockout technique.

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 6256
    Illegal EcoRI site found at 10357
    Illegal EcoRI site found at 11866
    Illegal XbaI site found at 9054
    Illegal SpeI site found at 2715
    Illegal PstI site found at 4500
    Illegal PstI site found at 11361
    Illegal PstI site found at 11608
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 6256
    Illegal EcoRI site found at 10357
    Illegal EcoRI site found at 11866
    Illegal NheI site found at 6015
    Illegal SpeI site found at 2715
    Illegal PstI site found at 4500
    Illegal PstI site found at 11361
    Illegal PstI site found at 11608
    Illegal NotI site found at 3034
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 6256
    Illegal EcoRI site found at 10357
    Illegal EcoRI site found at 11866
    Illegal BamHI site found at 8294
    Illegal BamHI site found at 10296
    Illegal XhoI site found at 4353
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 6256
    Illegal EcoRI site found at 10357
    Illegal EcoRI site found at 11866
    Illegal XbaI site found at 9054
    Illegal SpeI site found at 2715
    Illegal PstI site found at 4500
    Illegal PstI site found at 11361
    Illegal PstI site found at 11608
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 6256
    Illegal EcoRI site found at 10357
    Illegal EcoRI site found at 11866
    Illegal XbaI site found at 9054
    Illegal SpeI site found at 2715
    Illegal PstI site found at 4500
    Illegal PstI site found at 11361
    Illegal PstI site found at 11608
    Illegal AgeI site found at 10131
    Illegal AgeI site found at 11788
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI site found at 10113


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