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Multiplexed Gene Knock Out Cell Line Generation

Gene families or biological pathways usually contain several related genes that collaborate in the same role. Increasing demand in loss-of-function screens has warranted the need for a multiplexed gene knock out cell line. Multiplexing with CRISPR/Cas9 can accelerate genome engineering efforts in biopharmaceutical research and drug discovery.

Creative Biogene provides additional rounds of transformation and selection until every targeted gene is knocked out. Mammalian cells exposed to two or three gRNAs in turn resulted in the generation of cell lines harboring double or triple gene knockouts, that is, the successful disruption of two or three genes. All multiplexed knockout events were obtained with DNA mutations in targeted genes, and harbored the expected knockout phenotype(s). Investigation of entire gene families or biological pathways is enabled by our dual/triple gene knock out cell line generation service.

The KO status of each cell line is thoroughly tested as follows:

  • Biallelic mutation/deletion confirmed by PCR and DNA sequencing.
  • KO stability guaranteed up to 20 passages.
  • Both products enable unbiased, powerful loss-of-function screens.

Our services include:

  1. Pre-determined easy to transfect cell lines.
  2. Design and construct sgRNA vectors targeting to your genes.
  3. Transfect into cells.
  4. Clonal screening and selection.
  5. Additional rounds of transformation and selection.
  6. Validation of knockout by Sanger sequencing.
  7. Microbial/sterility testing


  • Targeted screening of specific pathways of importance.
  • Thorough screening of pathway or disease-focused genes.
  • Used as screening tools in the development of drugs.
  • To understand the mechanism of action of a drug.
  • To target specific biological processes.

Multiplexed Gene Knock Out Cell Line Generation Service Case

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