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Updated: Jan 10, 2026

Author Spotlight: Streamlining Rice Breeding with CRISPR/Cas for Obtaining Optimal Phenotypic and Agronomic Traits

Author Spotlight: Streamlining Rice Breeding with CRISPR/Cas for Obtaining Optimal Phenotypic and Agronomic Traits

Published on: January 3, 2025

TKC-MC: An Effective Strategy for Generating Heritable Heterozygous Mutations in Essential Genes in Rice.

Meilian Xu¹, Lang Yan^2,3, Min Zhu¹

¹National Key Laboratory of Crop Genetic Improvement and National Centre of Plant Gene Research (Wuhan), Hubei Hongshan Laboratory, Huazhong Agricultural University, Wuhan, China.

Plant Biotechnology Journal

|November 24, 2025

Summary

This summary is machine-generated.

Researchers developed Transgene-Killer CRISPR (TKC)-mediated mismatch-spacer targeting (TKC-M) to create heterozygous mutations in essential rice genes. This method overcomes CRISPR/Cas9

Keywords:

CRISPR/Cas9 TKC mismatch‐spacer rice transmission essential gene

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Identifying Mutations by High Resolution Melting in a TILLING Population of Rice

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Published on: September 2, 2019

Area of Science:

Plant genetics
Gene editing technologies
Molecular biology

Background:

CRISPR/Cas9 is vital for gene function studies and crop improvement.
Essential genes require heterozygous mutations for plant survival and propagation.
Standard CRISPR/Cas9 struggles to produce heterozygous mutants for essential genes in rice.

Purpose of the Study:

To develop an efficient method for generating heritable heterozygous mutations in essential rice genes.
To overcome limitations of traditional gene editing for essential gene research.

Main Methods:

Designed Transgene-Killer CRISPR (TKC)-mediated mismatch-spacer targeting (TKC-M).
Utilized a transgenic offspring self-elimination TKC platform with engineered gRNA-target mismatches.
Tested single and dual mismatches at specific gRNA positions for varying target sensitivities.
Employed a TKC-M Cocktail (TKC-MC) with multiple gRNA configurations.

Main Results:

TKC-M successfully enriched heritable heterozygous and mosaic incomplete-edited T0 mutants.
Single-base mismatches at positions 11 or 17 were effective for sensitive targets.
Dual mismatches at positions 8 and 15 maximized heterozygotes for insensitive targets.
TKC-MC significantly increased the incomplete-edited mutant ratio compared to standard TKC.

Conclusions:

TKC-M provides an efficient strategy for generating heterozygous mutations in essential rice genes.
This method facilitates research on essential genes and the creation of comprehensive mutant libraries.
Establishes a technical foundation for plant genome-wide mutant library generation.