CLICK: one-step generation of conditional knockout mice

BMC Genomics. 2018 May 2;19(1):318. doi: 10.1186/s12864-018-4713-y.


Background: CRISPR/Cas9 enables the targeting of genes in zygotes; however, efficient approaches to create loxP-flanked (floxed) alleles remain elusive.

Results: Here, we show that the electroporation of Cas9, two gRNAs, and long single-stranded DNA (lssDNA) into zygotes, termed CLICK (CRISPR with lssDNA inducing conditional knockout alleles), enables the quick generation of floxed alleles in mice and rats.

Conclusions: The high efficiency of CLICK provides homozygous knock-ins in oocytes carrying tissue-specific Cre, which allows the one-step generation of conditional knockouts in founder (F0) mice.

Keywords: CLICK; CRISPR/Cas9; Cre-loxP system; Long single-stranded DNA; Zygote electroporation.

MeSH terms

  • Alleles
  • Animals
  • Base Sequence
  • CRISPR-Cas Systems / genetics
  • Genetic Engineering / methods*
  • Injections
  • Mice
  • Mice, Knockout
  • Zygote / metabolism