籼稻垩白基因Chalk5功能标记引物设计与育种应用

doi:10.3969/j.issn.1006-8082.2026.05.013

摘要/Abstract

摘要：

垩白是衡量稻米外观品质和加工品质的关键指标之一。Chalk5是首个被克隆的、正向调控籼稻垩白率的主效基因。基于其功能性单核苷酸多态性（single nucleotide polymorphisms, SNPs）开发等位基因特异性PCR（allele-specific PCR，AS-PCR）标记，对于优化Chalk5在籼型优质稻分子标记辅助选择（marper-assisted selection, MAS）育种中的应用具有重要意义。本研究针对Chalk5基因启动子区的2个功能性SNP位点（-721和-485位），利用AS-PCR技术分别开发了共显性分子标记Chalk5_721和Chalk5_485，并对42份籼稻品种进行了基因型鉴定。在此基础上，结合Chalk5基因型、垩白表型及粒型性状开展了关联分析，旨在探讨该基因在籼型优质稻育种中的应用策略。结果表明，所设计的Chalk5功能标记能够通过一次常规PCR扩增及琼脂糖凝胶电泳，准确、快速地鉴别不同基因型。垩白度与Chalk5基因型呈显著相关性（卡方检验，P=0.04），其中Chalk5_721T和Chalk5_485T等位基因可显著降低籼稻垩白度。此外，在供试的42份籼稻材料中，垩白度与粒型长宽比呈极显著负相关（r=-0.8，P<0.001），表明细长粒型通常具有较低的垩白度。因此，在利用Chalk5功能标记进行优质稻选育时，应综合考量粒型、粒质量等产量相关性状。本研究为籼稻垩白性状的高效分子选择提供了实用工具与理论依据。

关键词: 籼稻, 垩白, Chalk5, 分子标记辅助选择, AS-PCR

Abstract:

Chalkiness is one of the key indicators for evaluating rice grain appearance quality and milling quality. Chalk5 is the first cloned major gene identified as positively regulating chalkiness rate in indica rice. Developing allele-specific PCR (AS-PCR) markers based on its functional single nucleotide polymorphisms (SNPs) is of great significance for optimizing the application of Chalk5 in marker-assisted selection (MAS) breeding for indica premium rice varieties. In this study, targeting two functional SNP loci (-721 and-485) located in the promoter region of the Chalk5 gene, we developed co-dominant molecular markers Chalk5_721 and Chalk5_485 using AS-PCR technology and conducted genotype identification for 42 indica rice accessions. Based on these data, association analysis was performed by integrating Chalk5 genotypes, chalkiness phenotypes, and grain shape traits to explore strategies for utilizing this gene in indica premium rice breeding. The results demonstrated that the designed Chalk5 functional markers could accurately and rapidly distinguish different genotypes through a single routine PCR amplification followed by agarose gel electrophoresis. Chalkiness degree showed a significant correlation with Chalk5 genotypes (Chi-square test, P=0.04), where the Chalk5_721T and Chalk5_485T alleles significantly reduced chalkiness in indica rice. Furthermore, in the tested panel of 42 indica rice materials, chalkiness degree exhibited a highly significant negative correlation with grain length-to-width ratio (r=-0.8, P<0.001), indicating that slender grains generally display lower chalkiness levels. Therefore, when applying Chalk5 functional markers for premium rice breeding, it is essential to comprehensively consider yield-related traits such as grain shape and grain weight. This study provides practical tools and theoretical support for the efficient molecular selection of chalkiness traits in indica rice.

Key words: indica rice, chalkiness, Chalk5, MAS, AS-PCR

中图分类号:

S511.0353

潘秀才, 陈波, 杜雪树, 房振兵, 赵沙沙, 孙强, 田永宏, 潘高峰. 籼稻垩白基因Chalk5功能标记引物设计与育种应用[J]. 中国稻米, 2026, 32(5): 74-81.

PAN Xiucai, CHEN Bo, DU Xueshu, FANG Zhenbing, ZHAO Shasha, SUN Qiang, TIAN Yonghong, PAN Gaofeng. Primer Design and Applications of Chalk5 Functional Markers for Indica Rice Breeding[J]. China Rice, 2026, 32(5): 74-81.

图/表 6

表1 42份籼稻品种Chalk5基因型鉴定和垩白度及粒型分析

表2 Chalk5基因AS-PCR引物序列

图1 Chalk5基因结构及引物位置示意图 A：Chalk5基因结构及2个功能性SNP位点；B：Chalk5_721和Chalk5_485标记引物结合位置；红色下划线核苷酸为功能SNP，蓝色小写核苷酸为引入错配碱基。

图2 Chalk5功能标记引物最佳退火温度筛选 A：Chalk5_721标记引物退火温度检测；B：Chalk5_485标记引物退火温度检测；M：DL2000分子量标记。

图3 Chalk5分子标记鉴定结果验证 A：9个具有不同Chalk5单倍型品种的琼脂糖凝胶电泳鉴定结果；B：7个具有纯合Chalk5单倍型品种的测序分析结果；C：2个杂合CT/AT型品种的测序峰图。

图4 部分籼稻品种的Chalk5基因型、垩白表型与粒型从左至右，稻米垩白度逐渐增高，粒型（长宽比）则逐渐降低；标尺为5 mm。

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