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Addgene

pSpCas9(BB)-2A-Puro (PX459) V2.0
(Plasmid #62988)

Ordering

This material is available to academics and nonprofits only.
Item Catalog # Description Quantity Price (USD)
Plasmid 62988 Standard format: Plasmid sent in bacteria as agar stab 1 $85
Cloning Grade DNA 62988-DNA.cg 2 µg of cloning grade DNA in Tris buffer 1 $105

Backbone

  • Vector backbone
    PX459
  • Total vector size (bp) 9200
  • Vector type
    Mammalian Expression, CRISPR
  • Selectable markers
    Puromycin

Growth in Bacteria

  • Bacterial Resistance(s)
    Ampicillin, 100 μg/mL
  • Growth Temperature
    37°C
  • Growth Strain(s)
    Stbl3
  • Copy number
    Unknown

Gene/Insert

  • Gene/Insert name
    hSpCas9-2A-Puro V2.0
  • Alt name
    SpCas9-2A-Puro V2.0
  • Alt name
    Cas9
  • Alt name
    PX459 V2.0
  • Species
    Synthetic
  • Insert Size (bp)
    6000
  • Promoter Cbh
  • Tags / Fusion Proteins
    • 3XFLAG (N terminal on insert)
    • 2A-Puro (C terminal on insert)

Cloning Information

  • Cloning method Restriction Enzyme
  • 5′ cloning site AgeI (not destroyed)
  • 3′ cloning site EcoRI (not destroyed)
  • 5′ sequencing primer TTTATGGCGAGGCGGCGG
  • 3′ sequencing primer puro-variant-R (5'-gtgggcttgtactcggtcat-3')
  • (Common Sequencing Primers)

Resource Information

Terms and Licenses

  • Academic/Nonprofit Terms
  • Industry Terms
    • Not Available to Industry
Trademarks:
  • Zeocin® is an InvivoGen trademark.

Depositor Comments

NOTE: This is an improved version of Addgene Plasmid #48139. It has improved puromycin selection. The EcoRI site is not unique and flanks the puromycin cassette.

For more information on Zhang Lab CRISPR Plasmids please refer to: http://www.addgene.org/crispr/zhang/

The guide sequence is cloned into this plasmid using BbsI sites. The hU6-F primer (5'-GAGGGCCTATTTCCCATGATT-3') can be used to confirm the gRNA sequence after cloning into the plasmid. Please see the cloning protocol provided by the Zhang lab for more details.

Information for Cloning Grade DNA (Catalog # 62988-DNA.cg) ( Back to top)

Purpose

Cloning grade DNA is suitable for use in PCR, cloning reactions, or transformation into E. coli. The purity and amount is not suitable for direct transfections.

Delivery

  • Amount 2 µg
  • Guaranteed Concentration 100 ng/µl +/- 5 ng/µl
  • Pricing $105 USD
  • Storage DNA can be stored at 4℃ (short term) or -20℃ (long term).

Terms and Licenses

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

Quality Control

Addgene has verified this plasmid using Next Generation Sequencing. Results are available here

How to cite this plasmid ( Back to top)

These plasmids were created by your colleagues. Please acknowledge the Principal Investigator, cite the article in which the plasmids were described, and include Addgene in the Materials and Methods of your future publications.

  • For your Materials & Methods section:

    pSpCas9(BB)-2A-Puro (PX459) V2.0 was a gift from Feng Zhang (Addgene plasmid # 62988 ; http://n2t.net/addgene:62988 ; RRID:Addgene_62988)
  • For your References section:

    Genome engineering using the CRISPR-Cas9 system. Ran FA, Hsu PD, Wright J, Agarwala V, Scott DA, Zhang F. Nat Protoc. 2013 Nov;8(11):2281-308. doi: 10.1038/nprot.2013.143. Epub 2013 Oct 24. 10.1038/nprot.2013.143 PubMed 24157548