pDB4283
(Plasmid #98702)

• Purpose: A plasmid containing the 3' portion of the bsdMX marker and the gRNA elements downstream of the target sequence. It serves as the PCR template for generating the gRNA insert 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: pEASY-blunt
• Backbone manufacturer: TransGen Biotech
• Vector type: Yeast Expression, CRISPR

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin and Kanamycin, 100 & 50 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: High Copy

Gene/Insert

• Gene/Insert name: gRNA PCR template
• Species: Synthetic

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.