Botanisches Institut

CRISPR/Cas Plasmids (S.pyogenes)

Gateway cloning plasmid set Streptococcus pyogenes

pDe-CAS9 (Nuclease) sequence information

  • Cas9 expression vector (Cas9 driven by the constitutive Ubi4-2 promotor from parsley)
  • Binary Gateway destination vector (contains ccdB gene!)
  • Bacterial selection: Spectinomycin (100 mg/l)
  • Plant selection: DL-Phosphinothricin (PPT, 6 mg/l)
    • BAR resistance cassette can easily be exchanged by HindIII
    • Please ask for pDe-CAS9-GentR if you are interested in gentamycin selection instead of PPT

 

pDe-CAS9-D10A (Nickase) sequence information

  • Cas9 nickase expression vector based on the pDe-CAS9 backbone
  • Plant selection: Kanamycin (30 mg/l)

 

pEn-Chimera (sgRNA) - sequence information

  • sgRNA coding vector (sgRNA can easily be customized for any target site of interest; sgRNA is driven by the Arabidopsis U6-26 promoter)
  • Gateway entry vector
  • Bacterial selection: Ampicillin (100 mg/l)

 

pEn-C1.1 (sgRNAs for paired nickases or paired nucleases) sequence information

  • sgRNA coding vector based on pEn-Chimera BUT allows cloning of up to 3 different sgRNAs into pDe-CAS9
  • Gateway entry vector
  • Bacterial selection: Ampicillin (100 mg/l)

 

Conventional cloning plasmid set Streptococcus pyogenes

pCAS9-TPC (Nuclease) sequence information

  • Cas9 expression vector (Cas9 driven by the constitutive Ubi4-2 promotor from parsley)
  • Bacterial selection: Spectinomycin (100 mg/l)
  • Plant selection: DL-Phosphinothricin (PPT, 6 mg/l)

 

pChimera (sgRNA) sequence information

  • sgRNA coding vector (sgRNA can easily be customized for any target site of interest; sgRNA is driven by the Arabidopsis U6-26 promoter)
  • sgRNA is not subcloned into pCAS9-TPC via Gateway cloning but via AvrII