pDB4281
(Plasmid #98700)

• Purpose: A Cas9-encoding plasmid containing the 5' portion of the bsdMX marker and the rrk1 promoter/leader. When linearized by NotI, it serves as the gapped vector in the split-bsdMX system
• Depositing Lab: Li-Lin Du
• Publication: Zhang et al G3 (Bethesda). 2018 Apr 27. pii: g3.118.200164. doi: 10.1534/g3.118.200164.

Backbone

• Vector backbone: pMZ374 (Addgene 59896)
• Vector type: Yeast Expression, CRISPR

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: High Copy

Gene/Insert

• Gene/Insert name: Cas9

Cloning Information

• Cloning method: Unknown

Terms and Licenses

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

Depositor Comments

These plasmids are used for a cloning-free method of CRISPR/Cas9-mediated genome editing in fission yeast. This method takes advantage of gap repair in fission yeast cells to assemble two linear DNA fragments, a gapped Cas9-encoding plasmid and a PCR-amplified gRNA insert, into a circular plasmid. To reduce non-specific gap repair background, we adopt a split-marker strategy. pDB4280 and pDB4282 serve as the source of the gapped plasmid and the PCR template for gRNA insert amplification, respectively, in the split-ura4 system. pDB4281 and pDB4283 serve the same purposes for the split-bsdMX system. pDB4279 is a transformation control for the split-bsdMX system.