Distribution of Grain Shape Related Genes in Rice Big Grain Germplasm BG1 and Elite Restorer Line Huazhan and Development of Relevant Functional Markers

doi:10.16819/j.1001-7216.2021.201211

Abstract

Abstract:

【Objective】The exploration and utilization of specific germplasm resources are one of the important means of rice breeding with high quality and high yield. Identifying the allelic variation types of some grain shape genes carried by Big Grain 1 (BG1) and Huazhan, a high quality restorer line widely used in rice breeding, and developing functional markers of corresponding genes are helpful to speed up the application of grain shape genes in rice breeding and improve the efficiency of accurate improvement of grain shape in rice.【Method】The target fragments of nine major grain shape genes (GS3, qLGY3, qGL3, GW2, GW8, GW5, TGW3, TGW6, GS9) in BG1 and Huazhan were sequenced and analyzed. According to the sequencing results of GW2, GS3, qGL3, TGW3, TGW6, qLGY3, GW8 and GW5, a functional marker for genotyping was developed.【Result】Compared with Nipponbare, 5 of the 9 major grain shape genes detected in BG1 showed functional deletion (gs3, GW8, gw2, tgw3, tgw6), 1 rare allele variation (qgl3) and 1 exon variable shear (qlgy3); Compared with Nipponbare, Huazhan had differences in 3 of the 9 major grain shape genes (gs3, GW8 and GW5). According to the sequencing results, the functional markers of GW2, GS3, QGL3, TGW3, TGW6, QLGY3, GW8 and GW5 were developed and 15 rice germplasm were used for detection. In the detection results, in addition to the marker tgw6-del of TGW6, which can only be used to identify whether there is allelic variation tgw6 of functional deletion type, the other 7 markers can accurately identify genotype. There was sequence variation of GS9 among the tested materials, which did not affect the change of grain shape. According to the above sequencing results, the functional markers of GW2, GS3, qGL3, TGW3, TGW6, qLGY3, GW8 and GW5 were developed and detected by 15 rice germplasms. Except for the marker TGW6-Del for TGW6, which can only be used to identify the allelic variant tgw6, a functional deletion type, the other seven markers can be genotyped accurately.【Conclusion】Compared with Nipponbare, the genes gs3, qgl3, qlgy3, tgw3, GW8 and tgw6 that regulate grain length in extra-large grain germplasm BG1 may have complex interaction pathways rather than simple effect accumulation, and the grain width may be the cumulative effect of gw2 and GW8. The slender grain shape of high quality restorer line Huazhan may be caused by the interaction effect of gs3, GW5 and GW8. The functional markers developed can be used in rice molecular marker-assisted breeding.

Key words: grain shape, grain length, grain width, grain weight, functional marker

摘要：

【目的】特异种质资源的发掘与利用是水稻优质高产育种的重要手段之一。明确特大籽粒种质BG1（Big Grain 1）和育种上广泛配组的优质恢复系华占携带的部分粒形基因等位变异类型,并开发相应基因的功能标记,有助于加快粒形基因的育种应用,提高水稻粒形精准改良的效率。【方法】对BG1和华占中的9个主效粒形基因（GS3、qLGY3、qGL3、GW2、GW8、GW5、TGW3、TGW6、GS9）的目的片段进行测序分析。并根据GW2、GS3、qGL3、TGW3、TGW6、qLGY3、GW8和GW5的测序结果,开发了鉴定其基因分型的功能标记。【结果】与日本晴相比,特大粒种质BG1在检测的9个主效粒形基因中有5个表现为功能缺失型（gs3、GW8、gw2、tgw3、tgw6）,1个稀有等位变异（qgl3）和1个外显子可变剪切型（qlgy3）;与日本晴相比,华占在检测的9个主效粒形基因中有3个（gs3、GW8、GW5）存在差异。根据测序结果开发了GW2、GS3、qGL3、TGW3、TGW6、qLGY3、GW8和GW5的功能标记,并利用15个水稻种质进行检测,检测结果中除了TGW6 的标记TGW6-Del 仅可用于鉴定是否含有功能缺失型的等位变异tgw6外,另外7个标记均为可准确鉴定基因型。【结论】与日本晴相比,特大粒种质BG1中调控粒长的基因gs3、qgl3、qlgy3、tgw3、GW8和tgw6之间可能存在复杂的互作调控通路,而不是简单的效应累加,粒宽特征可能是gw2、GW8的累加效应。优质恢复系华占的细长粒形可能是gs3、GW5和 GW8的互作效应所致。本研究中开发的功能标记可以用于水稻分子标记辅助育种。

关键词: 粒形, 粒长, 粒宽, 粒重, 功能标记

Ke GONG, Pao XUE, Xiaoxia WEN, Feifei LIAO, Bin SUN, Zequn PENG, Shihua CHENG, Liyong CAO, Yingxin ZHANG, Weixun WU, Lianping SUN, Xiaodeng ZHAN. Distribution of Grain Shape Related Genes in Rice Big Grain Germplasm BG1 and Elite Restorer Line Huazhan and Development of Relevant Functional Markers[J]. Chinese Journal OF Rice Science, 2021, 35(6): 543-553.

龚柯, 薛炮, 温小霞, 廖飞飞, 孙滨, 彭泽群, 程式华, 曹立勇, 张迎信, 吴玮勋, 孙廉平, 占小登. 水稻特大粒种质BG1和优质恢复系华占的粒形基因研究及相关功能标记开发[J]. 中国水稻科学, 2021, 35(6): 543-553.

Figures/Tables 10

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引物名称 Primer name	前引物 Forward primer	后引物 Reverse primer	产物大小 Product size/bp
GS3-seq	ACTGTATGCTCAAAGCATCT	TGCTTTACTTTTTTTTTTGC	1593
GL3.1-seq	GCCACTCATGCACCATAACTAC	ACCTCCTCGTAGACCTCCATAA	435
LGY3-cds-1	GCTGAAGCGGATCGAGAACAA	GCCGGATGGGATGTGTTCATT	744
LGY3-YZ	ACTTGTTACCACATCCAAAACGG	AATCCTGACAATAATTCGCCCA	522
GW2-cds-1	CAGGGACATCGACCAGAAGAA	TACAACCATGCCAACCCTTG	1199
GW5-indel-3	TCCATGCAGTACTTGGTTTCC	AGATCTGAGCGGTAACTTTCTC	938/2150
GW8-indel-2	AGCTACAGAATCCAGAAACAAACCA	CTGGCCCCTTCCCCTTGG	611
GS9-seq	GTTCCAACGCCAACAGCAC	CTTCGCGGTAGCGATCCAAT	2601
TGW3-cds-1	CCGGCTAGTTCGGTTGAATG	AGACGTTCGATGGCTTCTGG	1153/1233
TGW6-cds-1	CCACAGCCACAACGAGAATG	CGACTCCGACATACGGCAAT	947

引物名称 Primer name	前引物 Forward primer	后引物 Reverse primer	产物大小 Product size/bp
GS3-seq	ACTGTATGCTCAAAGCATCT	TGCTTTACTTTTTTTTTTGC	1593
GL3.1-seq	GCCACTCATGCACCATAACTAC	ACCTCCTCGTAGACCTCCATAA	435
LGY3-cds-1	GCTGAAGCGGATCGAGAACAA	GCCGGATGGGATGTGTTCATT	744
LGY3-YZ	ACTTGTTACCACATCCAAAACGG	AATCCTGACAATAATTCGCCCA	522
GW2-cds-1	CAGGGACATCGACCAGAAGAA	TACAACCATGCCAACCCTTG	1199
GW5-indel-3	TCCATGCAGTACTTGGTTTCC	AGATCTGAGCGGTAACTTTCTC	938/2150
GW8-indel-2	AGCTACAGAATCCAGAAACAAACCA	CTGGCCCCTTCCCCTTGG	611
GS9-seq	GTTCCAACGCCAACAGCAC	CTTCGCGGTAGCGATCCAAT	2601
TGW3-cds-1	CCGGCTAGTTCGGTTGAATG	AGACGTTCGATGGCTTCTGG	1153/1233
TGW6-cds-1	CCACAGCCACAACGAGAATG	CGACTCCGACATACGGCAAT	947

种质	千粒重	增幅	长宽比	增幅	粒长	增幅	粒宽	增幅
Germplasm	TGW/g	Increase/%	GL/GW	Increase/%	GL/mm	Increase/%	GW/mm	Increase/%
日本晴Nipponbare	26.11 ± 0.33		2.19 ± 0.01		6.98 ± 0.01		3.19 ± 0.01
BG1	62.97 ± 0.01	141.17	2.85 ± 0.04	30.14	12.18 ± 0.04	74.45	4.27 ± 0.06	33.86
华占Huazhan	18.44 ± 0.13	-29.38	3.85 ± 0.03	75.80	8.28 ± 0.08	16.62	2.15 ± 0.04	-32.60

种质	千粒重	增幅	长宽比	增幅	粒长	增幅	粒宽	增幅
Germplasm	TGW/g	Increase/%	GL/GW	Increase/%	GL/mm	Increase/%	GW/mm	Increase/%
日本晴Nipponbare	26.11 ± 0.33		2.19 ± 0.01		6.98 ± 0.01		3.19 ± 0.01
BG1	62.97 ± 0.01	141.17	2.85 ± 0.04	30.14	12.18 ± 0.04	74.45	4.27 ± 0.06	33.86
华占Huazhan	18.44 ± 0.13	-29.38	3.85 ± 0.03	75.80	8.28 ± 0.08	16.62	2.15 ± 0.04	-32.60

引物名称 Primer name	前引物 Forward primer	后引物 Reverse primer	内切酶 Enzyme	产物长度 Product size/bp
GS3-SNP	GGATCCACGCTGCCTCCAGATGGTG	TTGCCAAGGTTTTATAATCAATGGT	Hph Ⅰ	225/203
TGW3-SNP	CTACGATCCGTGGCCGAAA	TTACAATGAAAAAGCCACAGGCGTA	Rsa Ⅰ	206/185
qGL3-SNP	CGATTCTATCTGGTTCAGTGGTAGA	AAACAGGTTTTCTTACCTCCTCGT	Acc Ⅰ	212/192/170
GW2-InDel	CAACTCACACTGCTCAGCCTACA	ATACTCCACAGCATAACTGGGAGTCT	Pst Ⅰ	133/99
TGW6-InDel	AGCCCCAGCTACACGAAAAACAAGTCCGCG	CCCATTGGTGAAGCGAAGTG	Sac Ⅱ	221/196
LGY3-InDel	GGGCCTAATTTTGTCTTTGTTATTC	GTCTGCTGCTTCATTGCTCA	Sty Ⅰ	162/85/77
GW8-InDel	GCGTCAACACACAGCTCAAG	CGGCATCTTGAGATCCCACTC		89/79
GW5-InDel3	TCCATGCAGTACTTGGTTTCC	AGATCTGAGCGGTAACTTTCTC		2150/938