Journal of Zhejiang University-SCIENCE B

, Volume 19, Issue 8, pp 620–629 | Cite as

High-resolution melting-based TILLING of γ ray-induced mutations in rice

  • Shan Li
  • Song-mei Liu
  • Hao-wei Fu
  • Jian-zhong Huang
  • Qing-yao ShuEmail author


Targeting Induced Local Lesions IN Genomes (TILLING) is a reverse genetics strategy for the high-throughput screening of induced mutations. γ radiation, which often induces both insertion/deletion (Indel) and point mutations, has been widely used in mutation induction and crop breeding. The present study aimed to develop a simple, high-throughput TILLING system for screening γ ray-induced mutations using high-resolution melting (HRM) analysis. Pooled rice (Oryza sativa) samples mixed at a 1:7 ratio of Indel mutant to wild-type DNA could be distinguished from the wild-type controls by HRM analysis. Thus, an HRM-TILLING system that analyzes pooled samples of four M2 plants is recommended for screening γ ray-induced mutants in rice. For demonstration, a γ ray-mutagenized M2 rice population (n=4560) was screened for mutations in two genes, OsLCT1 and SPDT, using this HRM-TILLING system. Mutations including one single nucleotide substitution (G→A) and one single nucleotide insertion (A) were identified in OsLCT1, and one trinucleotide (TTC) deletion was identified in SPDT. These mutants can be used in rice breeding and genetic studies, and the findings are of importance for the application of γ ray mutagenesis to the breeding of rice and other seed crops.

Key words

Mutation screening High-resolution melting (HRM) analysis Targeting Induced Local Lesions IN Genomes (TILLING) Mutant Indel γ ray Rice 




建立适用于筛选伽马射线诱发突变的、基于高分辨率熔解曲线(high-resolution melting,HRM)技术的高通量定向诱导基因组局部突变技术 (Targeting Induced Local Lesions IN Genomes, TILLING)体系。




通过不同野生型/突变型比例混池DNA的HRM分析,确定HRM检测不同类型插入/缺失突变的能力,确定M2植株突变检测的适宜混池比例,并用一个伽玛诱变M2 群体(n=4560)筛选OsLCT1SPDT两个基因的突变体,确定实际效果。


以4 株M2植株混样,采用HRM可以有效检出突变。建立的基于HRM的TILLING体系适用于伽玛射线诱发突变的高通量筛选。


突变筛选 高分辨率熔解曲线(HRM) 定向诱导基因组局部突变技术(TILLING) 突变体 插入缺失 伽玛射线 水稻 

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Copyright information

© Zhejiang University and Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.National Key Laboratory of Rice Biology, Institute of Crop SciencesZhejiang UniversityHangzhouChina
  2. 2.Hubei Collaborative Innovation Center for Grain IndustryJingzhouChina
  3. 3.Institute of Nuclear Agricultural SciencesZhejiang UniversityHangzhouChina
  4. 4.Jiaxing Academy of Agricultural SciencesJiaxingChina

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