Genome Editing

CompoZr® Knockout ZFNs


Proven and Award Winning Zinc Finger Nuclease Technology

Sigma® Life Science offers ready-to-use, pre-constructed and functionally validated CompoZr Knockout Zinc Finger Nucleases (ZFNs) that are designed to permanently knockout any human, mouse or rat gene in a quick, simple and target specific manner. CompoZr Knockout ZFNs are a class of engineered DNA-binding proteins that facilitate highly specific targeted gene knockouts by creating a single double-strand break in a sequence-specific manner. CompoZr Knockout ZFNs are designed to a sequence within the first two-thirds of the open reading frame ensuring that the gene disruption will lead to a knockout. Upon receipt of ZFNs for your gene of interest, permanent and heritable knockout in any cell line where transfection is possible can be achieved much more rapidly and efficiently than with traditional knockout methods.


Ordering Information

Browse our catalog for a Knockout ZFN to any human, mouse or rat gene.

Browse Knockout ZFNs


For gene knockouts in other species visit the CompoZr Custom ZFN Service

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Benefits

Ready-To-Use

  • CompoZr Knockout ZFNs are designed, constructed and functionally validated before shipment
  • Receive ZFNs in plasmid or plasmid plus mRNA format
  • Immediately transfect your cell line once CompoZr Knockout ZFNs are received
  • Screen for gene disruption in as little as 48 hours
  • Single cell clone out individual cells with gene disruptions

Knockouts Induced through a Single Transfection Experiment

  • Single or biallelic edits occur in 1-20% of clone population
  • No antibiotic selection required for screening

High Specificity

  • CompoZr ZFN technology provides for targeted single locus gene modification
  • ZFN activity at non-specific sites is negligible (see references)
  • Genes disruptions at the target site are permanent and remain heritable

CompoZr Knockout ZFN Components

For increased access to ZFN technology Sigma Life Science provides Knockout ZFNs in two formats. The first format includes ZFNs as plasmids only, for an additional price ZFNs are provided as plasmids plus 10 aliquots of ready-to-use ZFN mRNA.

  • Plasmid DNA for each ZFN in the pair
  • Forward and reverse primers for screening the DNA locus for mutation
  • Genomic DNA as a positive control for PCR (from validated samples where the target gene has been mutated using the supplied CompoZr Knockout ZFN)
  • 10 aliquots of ready-to-deliver mRNA (optional component) 

Zinc Finger Nuclease References

Click here to view a list of references.



CompoZr® Knockout ZFNsFigure 1: CompoZr Knockout ZFNs

(A) Ready-to-transfect Zinc Finger Nucleases
ZFN pairs are provided as ready-to-transfect plasmid DNA and mRNA, for the cell line of your choice. Each format can be transfected into human cells and then translated into ZFN proteins.

 

(B) ZFN-induced double strand break
The ZFNs bind a specific DNA sequence within the target gene, inducing a double strand break at the bound site. This break is repaired by an imperfect endogenous repair mechanism called non-homologous end joining (NHEJ). Due to the imperfect nature of NHEJ, a percentage of double-strand breaks within a ZFN-treated cellular population will be misrepaired by the addition and/or deletion of nucleotides. These disruptions within the gene occur at the site of the break resulting in a non-sense gene product, the loss of gene function and a knockout. CompoZr Knockout ZFNs deliver highly successful gene knockout frequencies of up to 20% in several cell lines, without the need for any selection markers.

 

(C) Cel1 Assay
A nucleotide mismatch assay is used to detect the presence of mutant clones and quantify the effectiveness of ZFNs. The mismatch assay consists of amplifying the target region from ZFN-treated genomic DNA via standard PCR using the primers provided in the kit. Resultant PCR products are denatured and allowed to re-anneal. A fraction of re-annealed products will contain bulges of DNA where heterogeneous mismatches occur. When a mismatch sensitive enzyme is added to the reaction, DNA digestion will occur at the site of the nucleotide mismatch. PCR pools are analyzed via PAGE. Lanes 2 and 3 contain cells that have been edited via ZFN-induced DSBs. The presence of digested PCR product indicates ZFN-mediated mutagenesis and the ratio of digested PCR products to wildtype PCR product (quantified by densitometry) determines ZFN activity.




All Zinc Finger Nucleases are sold under license from Sangamo Biosciences. For a copy of the Label License provided with purchase of CompoZr ZFNs, please visit the ZFN Label License page. CompoZr is a trademark of Sigma-Aldrich Co. LLC

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