Cas9 (Streptococcus pyogenes CRISPR associated protein 9) is an endonuclease enzyme that, when recruited to a specific DNA sequence by the sgRNA (single guide RNA), introduces a double stranded break into the DNA. This double stranded break is repaired in mammalian cells either through Non-Homologous End Joining or Homologous Recombination. Non-Homologous End Joining often results in the deletion or insertion of several base pairs at the cut site, which, when resulting in a frameshift, causes the functional inactivation of the targeted gene. Cas9-expressing MCF7 cells can be transduced or electroporated with sgRNA targeting a gene of interest to quickly generate knock-out cell pools or cell lines.
Purchase of this cell line is for research purposes only; commercial use requires a separate license. View the full terms and conditions.
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Synonyms
SpCas9 cell pool, CRISPR cell pool, Cas9 cell pool, MCF7 cell pool
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Product Data Gallery
Flow cytometry analysis of intracellular expression of Cas9 in MCF7 cell pool
This cell line has been screened using the MycoAlert™ Mycoplasma Detection Kit (Lonza, #LT07-118) to confirm the absence of Mycoplasma contamination. MycoAlert Assay Control Set (Lonza, #LT07-518) was used as a positive control.
Storage/Stability
Cells will arrive upon dry ice and should immediately be thawed or stored in liquid nitrogen upon receipt. Do not use a -80°C freezer for long term storage.
Culture Conditions
Media Required for Cell Pool Culture
Thaw Medium 1 (BPS Bioscience #60187) + 10 μg/ml Insulin (Sigma-Aldrich #I0516): DMEM medium (Hyclone #SH30243.01) supplemented with 10% FBS (Life technologies #26140-079), and 1% Penicillin/Streptomycin (Hyclone #SV30010.01).
Growth Medium 1R (BPS Bioscience # 78180) + 10 μg/ml Insulin (Sigma-Aldrich #I0516): Thaw Medium 1 (BPS Bioscience, #60187) plus 0.25 μg/mL Puromycin (Invivogen, #ant-pr-1) to ensure recombinant expression.
Cells should be grown at 37°C with 5% CO2 using Growth Medium 1R plus 10 μg/ml Insulin (Sigma-Aldrich #I0516) to ensure recombinant expression is maintained.
Instructions for Use
See assay protocol for detailed instructions.
Applications
1. Quickly generating knock-out cell pools or cell lines in MCF7 cells. 2. Implementing sgRNA screens in Cas9-expressing MCF7 cells.
The CRISPR/CAS9 technology is covered under numerous patents, including U.S. Patent Nos. 8,697,359 and 8,771,945, as well as corresponding foreign patents applications, and patent rights.
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