1) Introduction:

CRE and FLP are site-specific recombinase. Each involves the recombination of sequences between short recognition target sites, LoxP for CRE, and FRT for FLP, allowing for precise manipulation of DNA sequence. They provides an excellent tool for conditional gene targeting.

Gentarget developed the GFP/RFP Color/Switch reporting lentivirus products, providing a very effective method for verifying the performance of CRE- or FLP-mediated recombination in vivo. (Notes: The optimal reaction temperature for Cre recombinase is 37°C, while for Flp recombinase it is 30°C . The CRE-LoxP has higher recombination efficiency than FLP-FRT.)

2) CRE or FLP recombination detection:

GenTarget ColorSwitch reporting lentivirus provide a convenient, easy, and fast indicator of CRE or FLP recombination efficacy in vivo or in cell culture. These lentivirus have been engineered to constitutively express the “LoxP-GFP-stop-LoxP-RFP-Stop” or “FRT-GFP-stop-FRT-RFP-Stop” cassette under either an enhanced CMV promoter or EF1a promoter. Those lentivirus detect the CRE or FLP enzyme efficiency via a color switch mechanism (see the expression cassette scheme below). 

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3) How it works:

This lentivirus demonstrates a strong GFP fluorescence after infection into any mammalian cells but does not show an RFP fluorescence signal (or some leaking RFP expression in the case of FLP reporting lentivirus). Once the CRE or FLP protein is present in the nucleus, the CRE or FLP excises / deletes the DNA fragment between two loxP sites or FRT sites, respectively ). As a result, the GFP is removed and RFP is expressed, resulted in high RFP expression. The RFP-expression % (or RFP-fluorescent intensity in the case for FLP) reflect the CRE (or FLP) recombination efficiency.  See the sample results below.

Images taken before and after the CRE expression Lentivirus (CAT#: LVP339-PBS) was added into cell culture (48~72 hours).

4) Product Features:

GenTarget provides the CRE and FLP reporting lentivirus with different antibiotic selection markers as puromycin, neomycin, or blasticidin. The selection markers are expressed under an RSV promoter (not shown in the scheme above). Therefore, you can easily select the positive transduced cells (expressing the CRE / FLP detection cassette) via a specific antibiotic marker, blasticidin (Bsd), Puromycin (puro), or Neomycin (Neo).

Please see Product manual (for CRE) and Product Manual (for FLP) details.

See also our Pre-made Color-Swtich Reporter cell lines that generated by transduction with those reporting lentivirus.