pKSEe401R
(Plasmid #218532)

• Purpose: (Empty Backbone) CRISPR/Cas9 genome editing vector pKSEe401R contains the maize codon-optimized Cas9 (zCas9) enhanced by the 5′-UTR fragment of OsMac3 and the pAtUBQ10-DsRED1-NOSt screening cassette in the backbone.
• Depositing Lab: Kirill Demchenko
• Publication: Kiryushkin et al Int J Mol Sci. 2024 Feb 6;25(4):1975. doi: 10.3390/ijms25041975.

Backbone

• Vector backbone: pKSE401
• Backbone manufacturer: Qi-Jun Chen (Addgene plasmid # 62202)
• Backbone size (bp): 18202
• Modifications to backbone: pKSEe401R contains the maize codon-optimized Cas9 (zCas9) enhanced by the 5′-UTR fragment of OsMac3 and the pAtUBQ10-DsRED1-NOSt screening cassette in the backbone.
• Vector type: Plant Expression
• Selectable markers: Neomycin (select with G418) ; pAtUBQ10-DsRED1

Growth in Bacteria

• Bacterial Resistance(s): Kanamycin and Spectinomycin, 50 & 50 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Growth instructions: Use kanamycin (50 mkg/ml) together with spectinomycin (50 mkg/ml) for empty plasmid propagation both in E. coli and in Agrobacterium strains. Spectinomycin allows saving the pressure against BsaI-containing cassette.
• Copy number: High Copy

Cloning Information

• Cloning method: Golden Gate
• 5′ sequencing primer: 35S prom: CTATCCTTCGCAAGACCCTTC; M13/pUC For: CCCAGTCACGACGTTGTAAAACG; DsRed1-N: GTACTGGAACTGGGGGGACAG
• 3′ sequencing primer: M13 Rev: CAGGAAACAGCTATGAC; DsRed1-C: AGCTGGACATCACCTCCCACAACG

Resource Information

• Supplemental Documents:
  • pKSEe401R.gb

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
• Industry Terms:
  • Not Available to Industry

Depositor Comments

As pKSEe401R is the modified version of original pKSE401 [Xing et al., 2014. BMC Plant Biology 14: 1-12], it allows to construct vectors expressing ONLY one gRNA for gene targeting. You may use original pCBC-DT1T2, pCBC-DT2T3, and pCBC-DT3T4 (AddGene IDs: 50590, 50591, 50592, respectively) for construction two-, three-, or four-gRNA-expressing vectors.