PUBLICATION

A PAGE screening approach for identifying CRISPR-Cas9-induced mutations in zebrafish

Authors
VanLeuven, A.J., Park, S., Menke, D.B., Lauderdale, J.D.
ID
ZDB-PUB-180626-6
Date
2018
Source
Biotechniques   64: 275-278 (Other)
Registered Authors
Lauderdale, James D.
Keywords
Danio rerio, genotyping, polymerase chain reaction (PCR)
MeSH Terms
  • Animals
  • CRISPR-Cas Systems/genetics*
  • DNA/analysis
  • DNA/genetics
  • Genotyping Techniques/methods*
  • Mutation/genetics*
  • Native Polyacrylamide Gel Electrophoresis/methods*
  • Polymerase Chain Reaction/methods*
  • Zebrafish/genetics*
PubMed
29939088 Full text @ Biotechniques
Abstract
The introduction of CRISPR-Cas9 technology for targeted mutagenesis has revolutionized reverse genetics and made genome editing a realistic option in many model organisms. One of the difficulties with this technique is screening for mutations in large numbers of samples. Many screening approaches for identifying CRISPR-Cas9 mutants have been published; however, in practice these methods are time consuming, expensive, or often yield false positives. This report describes a PCR-based screening approach using non-denaturing PAGE. This approach does not depend on the formation of heteroduplexes and reliably detects changes as small as 1 base-pair (bp) in nucleic acid length at the target site. This approach can be used to identify novel mutations and is also useful as a routine genotyping method.
Genes / Markers
Figures
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Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Antibodies
Orthology
Engineered Foreign Genes
Mapping