Mapping of qChalk8 Controlling Chalky Rice Rate in japonica Rice by Combining QTL-Seq with Molecular Markers

doi:10.16819/j.1001-7216.2024.231207

Abstract

Abstract:

【Objective】Chalkiness is an important characteristic that affects the appearance quality of rice grains. This study aims to use QTL-Seq and molecular marker mapping methods to locate QTLs related to chalky rice rates in japonica rice.【Method】F₂ segregating populations were constructed using the japonica germplasm Lamujia with a high chalky rice rate and the japonica variety Shuijing 3 with a low chalky rice rate. Two independent F₂ populations were planted in Yuanyang, Henan Province, and Sanya, Hainan Province, respectively. The single-plant chalky rice rate was examined, and extreme individuals were pooled for QTL-Seq analysis. Reliable QTLs were then subjected to verification using molecular marker mapping.【Result】QTL-Seq analysis of the two F₂ populations revealed the existence of a stable chalky rice rate QTL on chromosome 8, designated as qChalk8. Further molecular marker mapping confined the QTL to a genomic region between 17.6 Mb and 18.5 Mb. The LOD score for qChalk8 was 7.08, and qChalk8 could explain 15.9% of the phenotypic variation.【Conclusion】QTL-Seq and molecular marker mapping successfully identified qChalk8 related to chalky rice rates in japonica rice. The localization of qChalk8 establishes a foundation for further fine mapping and gene cloning of the chalkiness regulatory gene in japonica rice.

Key words: rice, grain chalkiness, chalky rice rate, QTL-Seq, QTL mapping

摘要：

【目的】垩白是影响稻米外观品质的重要性状。本研究旨在利用QTL-Seq和分子标记作图定位粳稻垩白粒率调控相关的QTL。【方法】利用高垩白粒率粳稻材料拉木加和低垩白粒率粳稻品种水晶3号构建F₂分离群体，将两份独立的F₂群体分别种植于河南原阳和海南三亚，两个群体单株垩白粒率考种后分别选取极端个体混池进行QTL-Seq分析，然后使用分子标记作图对稳定的QTL进行验证。【结果】两个F₂群体的QTL-Seq分析发现8号染色体存在一个较为稳定的QTL位点qChalk8。进一步通过分子标记作图将该QTL定位于17.6－18.5 Mb，该QTL位点的LOD值为7.08，表型贡献率为15.9%。【结论】利用QTL-Seq和分子标记作图定位了粳稻垩白粒率相关QTL-qChalk8，为粳稻垩白调控基因进一步的精细定位和基因克隆奠定了基础。

关键词: 水稻, 垩白, 垩白粒率, QTL-Seq, QTL定位

DU Yanxiu, SUN Wenyu, YUAN Zeke, ZHANG Qianqian, LI Fuhao, LI Junzhou, SUN Hongzheng. Mapping of qChalk8 Controlling Chalky Rice Rate in japonica Rice by Combining QTL-Seq with Molecular Markers[J]. Chinese Journal OF Rice Science, 2024, 38(6): 665-671.

杜彦修, 孙文玉, 袁泽科, 张倩倩, 李富豪, 李俊周, 孙红正. 利用QTL-Seq结合分子标记定位粳稻垩白粒率控制位点qChalk8[J]. 中国水稻科学, 2024, 38(6): 665-671.

Figures/Tables 6

Table 1. Primers used in molecular marker mapping in this study

引物名称Primer	位置 Location	正向序列 Forward sequence	反向序列 Reverse sequence
8_0.8Mb	Chr8_837961	CCGGCATTTTGACAGGAAGTG	ACCTCTCTCTCTTTCTCTCTCCC
8_3.5Mb	Chr8_3532447	TCCAGATTCGTGATATTAGGATGAGT	GGAGGATCAGAGAGACCATGC
8_4.5Mb	Chr8_4537532	TCGTCGGGATTATTCCGAGC	ACACCGCCACCAATCAAAGA
8_7.7Mb	Chr8_7747072	CGGTCCATTGCACACCCTTA	ACATAACACATGCACCATGGA
8_9.4Mb	Chr8_9385739	TTGATGGGAGTGCCTGAAGA	TCACTGGTTACTATTGGTCACGT
8_15.8Mb	Chr8_15858062	GTAAGAAGCCAGAGCCGGAG	CTAAGAGAAGAGCGGGGTGC
8_16.5Mb	Chr8_16560856	ACAACCAACCCGGCTTGTTA	ACCATCCATTTGGCCTTGTT
8_16.9Mb	Chr8_16906251	CGACGAGGGGTCTTACCAAG	CTTCTCCCGATCCGCTCAAC
8_17.6Mb	Chr8_17683064	ACTTGCAACAACGATTGACCT	AAGCCACCAACCTTATTGCA
8_18.5Mb	Chr8_18507050	TGCTTTTCGCTTGGGCTAGA	GTTCCACTCTGCTACGGCTT
8_19.6Mb	Chr8_19583199	TAAACCCTTCAGTCCACGGC	ATGAGCTGGGGTATCGACCT
8_21.8Mb	Chr8_21807790	TCGCTCGCTTCCTGAAGTTT	GAAAAGCTCTGGCCAAACCG

Table 2. Measurement of chalkiness rate and grain shape traits of Shuijing 3 and Lamujia

性状 Trait	水晶3号 Shuijing 3	拉木加 Lamujia
垩白粒率 Chalky rice rate (%)	8.1±0.5	91.9±0.3
粒长Grain length (mm)	4.8±0.0	4.9±0.0
粒宽 Grain width (mm)	2.7±0.0	2.8±0.0
长宽比 Length-width ratio	1.8±0.0	1.7±0.0

Fig. 1. Chalky rice rates of Shuijing 3, Lamujia and their reciprocal cross F1 in two environments A, Chalky rice rate of Shuijing 3 and Lamujia in Sanya, Hainan Province and Yuanyang, Henan Province; B, Chalky rice rate of reciprocal cross F1 in Sanya and Yuanyang. SLF1 is Shuijing 3×Lamujia F1 and LSF1 is Lamujia×Shuijing 3 F1.

Fig. 2. Histogram of chalky rice rate distribution of Hainan and Henan F2 populations A, F2 population of Sanya, Hainan; B, F2 population of Yuanyang, Henan. The dashed line rectangles indicate high chalkiness and low chalkiness bulks.

Fig. 3. ΔSNP-index analysis of extreme individuals bulks of Hainan and Henan F2 populations A, ΔSNP-index of extreme individuals bulks of Hainan F2 population, the red line indicates 95% confidence interval and the blue line indicates 99% confidence interval; B, ΔSNP-index of extreme individuals bulks of Henan F2 population; C, SNP-index of high chalkiness bulk (blue dots) and low chalkiness bulk (green) in Hainan population; D, SNP-index of high chalkiness bulk (blue dots) and low chalkiness bulk (green) in Henan population.

Fig. 4. Genetic linkage map of chalky rice rate and molecular markers on chromosome 8

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