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Related Experiment Videos

A simple and efficient method for CRISPR/Cas9-induced mutant screening.

Yufeng Hua1, Chun Wang1, Jian Huang2

  • 1State Key Laboratory of Rice Biology, China National Rice Research Institute, Chinese Academy of Agricultural Sciences, Hangzhou 310006, China.

Journal of Genetics and Genomics = Yi Chuan Xue Bao
|April 19, 2017
PubMed
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We developed a cost-effective annealing at critical temperature PCR (ACT-PCR) method for screening CRISPR/Cas9-induced mutants. This simple technique accurately identifies mutants in plants and animals, offering a faster alternative to existing methods.

Area of Science:

  • Molecular Biology
  • Genetics
  • Biotechnology

Background:

  • The CRISPR/Cas9 system is a powerful tool for generating genetic mutations.
  • Existing methods for screening CRISPR/Cas9-induced mutants are often time-consuming, labor-intensive, and costly.
  • There is a need for a more efficient and accessible screening technique.

Purpose of the Study:

  • To develop and validate a cost-effective and sensitive screening method for CRISPR/Cas9-induced mutants.
  • To provide a simpler alternative to existing mutant screening assays.
  • To enable rapid, large-scale screening of mutants in various organisms.

Main Methods:

  • Developed a novel screening technique based on conventional PCR called annealing at critical temperature PCR (ACT-PCR).
Keywords:
ACT-PCRCRISPR/Cas9Genome editingMutant screening

Related Experiment Videos

  • ACT-PCR involves a single PCR step followed by agarose gel electrophoresis.
  • Validated the method in rice and zebrafish, and for cultured cells.

    • Main Results:

    • ACT-PCR accurately distinguished CRISPR/Cas9-induced mutants from wild-type samples.
    • The method demonstrated sensitivity and cost-effectiveness compared to traditional assays.
    • Successfully adapted ACT-PCR for determining mutation frequency in cultured cells.

    Conclusions:

    • ACT-PCR is a simple, cost-effective, and sensitive method for screening CRISPR/Cas9-induced mutants.
    • This technique is suitable for rapid, large-scale mutant screening in both plants and animals.
    • ACT-PCR offers a valuable alternative for researchers working with CRISPR/Cas9 technology.