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Chromovert® Technology: Chromo-Tag™ Probes and Plasmids

CHROMOVERT TECHNOLOGY is a newly published research tool for rapid creation of stable cell lines. The technology utilizes fluorogenic oligonucleotide signaling probes and flow cytometry to detect and isolate individual living cells expressing one or more genes.

ABOUT CHROMOVERT® TECHNOLOGY

Mammalian cell lines, especially those produced using immortalized lines like human embryonic kidney 293 (HEK 293) and Chinese hamster ovary (CHO) cells, are widely used in biological research, including in drug discovery and for biologics production. In general, cell line production begins with the introduction of one or more plasmids encoding cDNAs of interest into a cell culture. The goal is to produce, in a reasonable time frame, clonal cell lines that meet desired criteria for an application of interest. Despite multiple advances in cell engineering, the rapid creation of robust and multi-gene cell lines remains of reach - until now.

Originating at The Rockefeller University, Chromovert Technology is a cell engineering tool for rapid production of stable cell lines expressing one or more genes. The method is based upon the broadly-applicable principles of fluorescence-resonance energy transfer (FRET) and nucleic acid hybridization using fluorogenic oligonucleotide signaling probes originally reported for in tubo qRT-PCR applications, transfected into living cells. The termini of the signaling probes are covalently linked to a fluorophore or quencher paired to absorb its emission. The termini are designed to form a 4-7 base-pair stem juxtaposing the fluorophore and quencher pair. In the presence of target sequence, hybridization of the sequence-specific probe results in a fluorogenic conformational change. Flow cytometry is then used to detect and isolate positive cells that fluoresce above background. Thousands of individual clones can then be isolated and expanded using automated cell culture methods. Functional testing over time in the absence of selective pressure is used to select final clones.

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CHROMOVERT® PROCESS
a) cDNAs are subcloned for expression of mRNAs comprising 3' untranslated plasmid-encoded Chromo-Tag™ sequences for detection using fluorogenic oligonucleotide signaling probes. Protein expression products remain untagged.
b) To create cell lines, one or more Chromo-Tagged cDNAs are transfected into cells, the transfected cells are exposed to differentially-labeled signaling probes and individual positive cells are isolated using flow cytometry. Downstream testing is used to select final cell lines.

CHROMOVERT® PUBLICATION
Cell engineering method using fluorogenic oligonucleotide signaling probes and flow cytometry. Shekdar, K., Langer, J., Venkatachalan, S. et al. Cell engineering method using luorogenic oligonucleotide signaling probes and flow cytometry. Biotechnol Lett (2021). https://doi.org/10.1007/s10529-021-03101-5


For more information and for complete pChromo-Plasmid™ sequences, go to Secondcell Bio, LLC at https://www.secondcellbio.com/.

Click here to order Chromo-Tag™ Probes and Plasmids.

RESEARCH-USE ONLY

RESEARCH MATERIALS, INCLUDING FLUOROGENIC PROBES AND PCHROMO-PLASMIDS™, COMPRISING CHROMO-TAGS™, ARE LICENSED FOR RESEARCH PURPOSES ONLY. AN INDIVIDUAL LICENSE FOR COMMERCIAL USE MAY BE OBTAINED BY CONTACTING SECONDCELL BIO, LLC AT CELLS@SECONDCELLBIO.COM. GENE LINK, INC. IN COLLABORATION WITH SECONDCELL BIO, LLC, IS MAKING THESE PRODUCTS AVAILABLE FOR RESEARCH USE ONLY WITHOUT FURTHER LICENSING.
This product is provided under an intellectual property license from Secondcell Bio, Inc. The transfer of this product is conditioned on the buyer using the purchased product solely in research conducted by the buyer, excluding contract research or any fee for service research, and the buyer must not: (1) use this product or its components for (a) diagnostic, therapeutic or prophylactic purposes, (b) testing, analysis, or screening services or information in return for compensation on a per-test basis, or (c) manufacturing or quality assurance or quality control; and/or (2) sell or transfer this product or its components for resale, whether or not resold for use in research. The buyer acknowledges and agrees that it is not entitled to any service or support with respect to this product. For information on purchasing a license to this product for purposes other than as described above, or potential service or support, contact Secondcell Bio, LLC at 40 Remsen Street, #4, Brooklyn, N.Y. 11201 or cells@secondcellbio.com.

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