Genome Editing

CompoZr® Zinc Finger Nucleases FP-ZFN-IDLV
(Integration-Deficient Lentivirus)

 Deliver Knockout ZFNs or Custom ZFNs with Fluorescent Proteins via IDLV

CompoZr Zinc Finger Nucleases (ZFNs) are a class of engineered DNA-binding proteins that facilitate highly specific targeted editing of any genome by creating double-stranded breaks in DNA at user specified locations. You can use ZFNs to create your choice of stable genetically engineered cell lines or organisms with gene deletions, gene integrations and gene modifications.

The following graphic highlights the abundance of genetic modifications that may be conducted using CompoZr ZFNs. To learn more about CompoZr ZFNs, please visit: sigma.com/zfns.

The new Sigma CompoZr FP-ZFN-IDLV products provide researchers with a lentiviral delivery format to increase the efficiency of genome modifications in various cell types, including hard-to-transfect cells. To preserve the transient mode of ZFN expression typically implemented in genome editing workflows, we use a system that enables integration-deficient lentiviral (IDLV) packaging to minimize integration of ZFN transgenes into the host genome.

To address expression limitations, we have constructed vectors that link ZFN expression directly to fluorescent protein reporters. This reporter-linked expression format enables enrichment of cell populations that have undergone both efficient transduction and subsequent high level expression of ZFN transgenes and stand to substantially increase genome editing frequencies.

Product No. Description Pricing
CSTZV1
CompoZr Custom ZFN IDLV Plasmid Kit
Check pricing
CSTZV2 CompoZr Custom ZFN IDLV Plasmid and Virus Kit Check pricing
ZVCTRLD CompoZr ZFN FP-IDLV Non-Targeting Control Plasmid DNA Check pricing
ZVCTRLV CompoZr ZFN FP-IDLV Non-Targeting Control Check pricing
I3411 IDLV Packaging Mix Check pricing


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 Benefits

  • Combines CompoZr FP-ZFNs with a lentiviral delivery system
  • Use of FP-ZFNs enable enrichment of ZFN activity via FACS
  • Preserves transient mode of ZFN expression using IDLV packaging – avoids permanent integration of ZFN transgenes into the genome
  • IDLV gene expression of ZFNs enables manipulation of primary cells, non-dividing cells and a variety of cell types.

Figure 1. The FP-ZFN-IDLV vector allows more rapid and accurate assessment of the performance of lentiviral ZFN expression control elements. This reporter-linked expression format enables enrichment of cell populations and can substantially increase genome editing frequencies in clonal cell populations.

 

Figure 2. Schematic of fluorescent protein-linked ZFNs (left). GFP is linked to the N-terminus of left ZFN (ZF-FokI-ELD) and RFP to the N-terminus of right ZFN (ZF-FokI-KKR) by a 2A peptide sequence. ZFN expression cassette activity can be rapidly monitored post-nucleofection via microscopy (K562 cells – image on the right).

 

Figure 3. MSC cells are hard to transfect cells that when nucleofected with ZFNs in plasmid format they undergo complete apoptosis within 24 hours. A) MSC cells show good viability and visible modification when treated with ZFNs in IDLV format. B) MSC cells were pooled sorted (upper gate), and ZFN cutting efficiency determined by the CEL-I assay (C). The CEL-I assay is a nucleotide mismatch assay used to detect the presence of mutant clones and quantify the effectiveness of ZFNs. The detected cutting efficiency of ZFNs was 9 times greater in the high sort cells compared to pooled cells. The CEL-I uncut wild-type band is 398 bp, and the ZFN modified bands are 234 bp and 164 bp.