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Methods Mol Biol. 2017;1679:169-185. doi: 10.1007/978-1-4939-7337-8_11.

Targeted Mutagenesis in Hexaploid Bread Wheat Using the TALEN and CRISPR/Cas Systems.

Author information

1
State Key Laboratory of Plant Cell and Chromosome Engineering, and Center for Genome Editing, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, China.
2
University of Chinese Academy of Sciences, Beijing, 100049, China.
3
State Key Laboratory of Plant Cell and Chromosome Engineering, and Center for Genome Editing, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, China. cxgao@genetics.ac.cn.

Abstract

The use of sequence-specific transcription activator-like effector nucleases (TALENs) and the clustered regularly interspaced short palindromic repeats-associated system (CRISPR/Cas9) have provided powerful reverse genetic approaches to the targeted modification of genomes in numerous organisms. Both systems have been employed to generate loss-of-function alleles in bread wheat, by targeting multiple and single copies of genes. Here we present protocols for modifying the wheat genome using the two systems. The protocols include the design of TALEN and CRISPR/Cas9 target sites and their construction, evaluation of their activities in protoplasts, transformation of plants, and mutation screening.

KEYWORDS:

CRISPR/Cas9; Genome editing; TALEN; Targeted mutagenesis; Wheat; Wheat protoplasts

PMID:
28913801
DOI:
10.1007/978-1-4939-7337-8_11
[Indexed for MEDLINE]

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