pML104
(Plasmid #67638)

• Purpose: Expresses Cas9 and contains guide RNA expression cassette with BclI-SwaI cloning sites for guide sequence cloning; Contains URA3 marker for yeast transformation
• Depositing Lab: John Wyrick
• Publication: Laughery et al Yeast. 2015 Dec;32(12):711-20. doi: 10.1002/yea.3098. Epub 2015 Sep 21.

Backbone

• Vector backbone: pRSII426
• Backbone size w/o insert (bp): 5718
• Total vector size (bp): 11240
• Vector type: Yeast Expression, CRISPR
• Selectable markers: URA3

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Growth instructions: Need to purify DNA from Dam- E.coli strain for efficient BclI cutting
• Copy number: High Copy

Gene/Insert 1

• Gene/Insert name: Cas9
• Species: S. cerevisiae (budding yeast); Streptococcus pyogenes
• Insert Size (bp): 4151
• Promoter: pTDH3

Cloning Information for Gene/Insert 1

• Cloning method: Restriction Enzyme
• 5′ cloning site: EcoRI (not destroyed)
• 3′ cloning site: XhoI (not destroyed)
• 5′ sequencing primer: T7

Gene/Insert 2

• Gene/Insert name: single guide RNA expression cassette
• Species: S. cerevisiae (budding yeast), Synthetic
• Insert Size (bp): 447
• Promoter: pSNR52

Cloning Information for Gene/Insert 2

• Cloning method: Restriction Enzyme
• 5′ cloning site: SacII (not destroyed)
• 3′ cloning site: EagI (not destroyed)
• 5′ sequencing primer: T3

Resource Information

• A portion of this plasmid was derived from a plasmid made by: Cas9 gene cloned from pTDH3-dCas9 and mutated extensively
• Articles Citing this Plasmid:
  • 21 References

Terms and Licenses

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