基于PARMS技术的水稻粒形基因GW8分子标记的开发
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摘要
【目的】开发水稻粒宽调控基因GW8的荧光分子标记,为快速、准确鉴定其基因型提供技术支持。【方法】通过与宽粒亲本GW8基因序列进行比对可知,窄粒杂交稻亲本美B的GW8基因序列在第二内含子有2个碱基缺失差异的特性之一,据此结合五引物扩增受阻突变体系技术,建立GW8基因荧光分子标记PM-GW8。【结果】利用荧光分子标记PM-GW8对42份籼稻亲本材料进行鉴定,仅在美B中检测到有2个碱基缺失的荧光信号。对亲本材料的谷粒宽度进行测量,其中美B的粒宽相对其他品种较窄,验证了该分子标记检测GW8的基因型与粒宽表型相符合。【结论】GW8荧光分子标记PM-GW8能快速检测水稻是否存在2个碱基缺失的GW8基因,为利用分子标记辅助选择改良水稻粒宽提供高效、可靠的基因分型技术。
【Objective】In order to provide technical support for efficient and accurate genotyping, the fluorescence molecular markers for rice grain width regulation gene GW8 was developed.【Method】Based on one of the characters of two bases deletion on the second intron of GW8 gene in narrow grain hybrid rice parent MeiB by comparison with wide gain varieties, and combined with penta-primer amplification refractory mutation system, the fluorescence molecular marker PM-GW8 of GW8 gene was developed.【Result】42 indica rice varieties were tested by the PM-GW8 marker, data showed that fluorescence signal of 2 bases deletion of GW8 was only detected in MeiB. These varieties were also analysis with grain width measurement, grain of MeiB was more narrow than grain of other varieties, this result indicated that genotype of GW8 detected by the marker was corresponding to phenotype of grain width.【Conclusion】The fluorescence molecular marker PM-GW8 can be used to detect whether two bases deletion in GW8 gene exist in rice rapidly, providing an efficient and reliable gene screening technic for the use of marker-assisted approach to improve rice gain width.
【Objective】In order to provide technical support for efficient and accurate genotyping, the fluorescence molecular markers for rice grain width regulation gene GW8 was developed.【Method】Based on one of the characters of two bases deletion on the second intron of GW8 gene in narrow grain hybrid rice parent MeiB by comparison with wide gain varieties, and combined with penta-primer amplification refractory mutation system, the fluorescence molecular marker PM-GW8 of GW8 gene was developed.【Result】42 indica rice varieties were tested by the PM-GW8 marker, data showed that fluorescence signal of 2 bases deletion of GW8 was only detected in MeiB. These varieties were also analysis with grain width measurement, grain of MeiB was more narrow than grain of other varieties, this result indicated that genotype of GW8 detected by the marker was corresponding to phenotype of grain width.【Conclusion】The fluorescence molecular marker PM-GW8 can be used to detect whether two bases deletion in GW8 gene exist in rice rapidly, providing an efficient and reliable gene screening technic for the use of marker-assisted approach to improve rice gain width.
引文
[1]Sakamoto T, Matsuoka M. Identifying and exploiting grain yield genes in rice[J]. Current Opinion in Plant Biology, 2008, 11(2): 209-214.
[2]Wang S K, Wu K, Yuan Q B, et al. Control of grain size, shape and quality by OsSPL16 in rice[J]. Nature Genetics, 2012, 44(8): 950-954.
[3]刘喜, 牟昌铃, 周春雷, 等. 水稻粒型基因克隆和调控机制研究进展[J]. 中国水稻科学, 2018, 32(1): 1-11.
[4]Song X J, Huang W, Shi M, et al. A QTL for rice grain width and weight encodes a previously unknown RING-type E3 ubiquitin ligase[J]. Nature Genetics, 2007, 39(5): 623-630.
[5]Liu J, Chen J, Zheng X, et al. GW5 acts in the brassinosteroid signalling pathway to regulate grain width and weight in rice[J]. Nature Plants, 2017(3): 1-7.
[6]Weng J, Gu S, Wan X, et al. Isolation and initial characterization of GW5, a major QTL associated with rice grain width and weight[J]. Cell Research, 2008, 18(12): 1199-1209.
[7]Wan X, Weng J, Zhai H, et al. Quantitative trait loci (QTL) analysis for rice grain width and fine mapping of an identified QTL allele gw-5 in a recombination hotspot region on chromosome 5[J]. Genetics, 2008, 179(4): 2239-2252.
[8]Ishimaru K, Hirotsu N, Madoka Y, et al. Loss of function of the IAA-glucose hydrolase gene TGW6 enhances rice grain weight and increases yield[J]. Nature Genetics, 2013, 45(6): 707-711.
[9]Wang Y, Xiong G, Hu J, et al. Copy number variation at the GL7 locus contributes to grain size diversity in rice[J]. Nature Genetics, 2015, 47(8): 944-948.
[10]Wang S, Li S, Liu Q, et al. The OsSPL16-GW7 regulatory module determines grain shape and simultaneously improves rice yield and grain quality[J]. Nature Genetics, 2015, 47(8): 949-954.
[11]Zhou Y, Miao J, Gu H, et al. Natural variations in SLG7 regulate grain shape in rice[J]. Genetics, 2015, 201(4): 1591-1599.
[12]Fan C, Xing Y, Mao H, et al. GS3, a major QTL for grain length and weight and minor QTL for grain width and thickness in rice, encodes a putative transmembrane protein[J]. Tag. Theoretical & Applied Genetics. Theoretische Und Angewandte Genetik, 2006, 112(6): 1164-1171.
[13]Hu Z, He H, Zhang S,et al. A kelch motif-containing serine/threonine protein phosphatase determines the large grain QTL trait in rice[J]. Journal of Integrative Plant Biology, 2012, 54(12): 979-990.
[14]Qi P, Lin Y S, Song X J, et al. The novel quantitative trait locus GL3.1 controls rice grain size and yield by regulating Cyclin-T1;3[J]. Cell Research, 2012, 22(12): 1666-1680.
[15]Zhang X, Wang J, Huang J, et al. Rare allele of OsPPKL1 associated with grain length causes extra-large grain and a significant yield increase in rice[J]. Proceedings of the National Academy of Sciences of the United States of America, 2012, 109(52): 21534-21539.
[16]Li Y, Fan C, Xing Y, et al. Natural variation in GS5 plays an important role in regulating grain size and yield in rice[J]. Nature Genetics, 2011, 43(12): 1266-1269.
[17]Xu C, Liu Y, Li Y, et al. Differential expression of GS5 regulates grain size in rice[J]. Journal of Experimental Botany, 2015, 66(9): 2611-2623.
[18]Sun L, Li X, Fu Y, et al. GS6, a member of the GRAS gene family, negatively regulates grain size in rice[J]. Journal of Integrative Plant Biology, 2013, 55(10): 938-949.
[19]Kesavan M,Song J T,Seo H S. Seed size: a priority trait in cereal crops[J]. Physiologia Plantarum, 2013, 147(2): 113-120.
[20]Li N and Li Y H. Signaling pathways of seed size control in plants[J]. Current Opinion in Plant Biology, 2016, 33: 23-32.
[21]裔传灯, 王德荣, 蒋伟, 等. 水稻粒形基因GS5的功能标记开发和单体型鉴定[J]. 中国水稻科学, 2016, 30(5): 487-492.
[22]裔传灯, 李玮, 王德荣, 等. 水稻粒形基因GS3的功能标记开发与鉴定[J]. 江苏农业科学,2016, 44(12): 64-67.
[23]裔传灯, 王德荣, 蒋伟, 等. 水稻粒形基因GW8的功能标记开发和单体型鉴定[J]. 作物学报, 2016, 42(9): 1291-1297.
[24]Ye S, Dhillon S, Ke X, et al. An efficient procedure for genotyping single nucleotide polymorphisms[J]. Nucleic Acids Research, 2001, 29(17): e88.
[25]卿冬进, 刘开强, 杨燕宇, 等. 基于PARMS技术的抗稻瘟病基因Pigm分子标记的开发[J]. 西南农业学报, 2018, 31(8): 1617-1621.
[26]Murray M G,Thompson W F. Rapid isolation of high molecular weight plant DNA[J]. Nucleic Acids Research, 1980, 8(19): 4321.
[27]李欣, 莫惠栋, 王安民, 等. 粳型杂种稻米品质性状的遗传表达[J]. 中国水稻科学, 1999, 13(4): 197-204.
[28]殷得所, 夏明元, 李进波, 等. 抗稻瘟病基因Pi9的STS连锁标记开发及在分子标记辅助育种中的应用[J]. 中国水稻科学, 2011, 25(1): 25-30.
[29]Huang K, Wang D, Duan P, et al. WIDE AND THICK GRAIN 1, which encodes an otubain-like protease with deubiquitination activity, influences grain size and shape in rice[J]. Plant Journal, 2017, 91(5): 849-860.
[2]Wang S K, Wu K, Yuan Q B, et al. Control of grain size, shape and quality by OsSPL16 in rice[J]. Nature Genetics, 2012, 44(8): 950-954.
[3]刘喜, 牟昌铃, 周春雷, 等. 水稻粒型基因克隆和调控机制研究进展[J]. 中国水稻科学, 2018, 32(1): 1-11.
[4]Song X J, Huang W, Shi M, et al. A QTL for rice grain width and weight encodes a previously unknown RING-type E3 ubiquitin ligase[J]. Nature Genetics, 2007, 39(5): 623-630.
[5]Liu J, Chen J, Zheng X, et al. GW5 acts in the brassinosteroid signalling pathway to regulate grain width and weight in rice[J]. Nature Plants, 2017(3): 1-7.
[6]Weng J, Gu S, Wan X, et al. Isolation and initial characterization of GW5, a major QTL associated with rice grain width and weight[J]. Cell Research, 2008, 18(12): 1199-1209.
[7]Wan X, Weng J, Zhai H, et al. Quantitative trait loci (QTL) analysis for rice grain width and fine mapping of an identified QTL allele gw-5 in a recombination hotspot region on chromosome 5[J]. Genetics, 2008, 179(4): 2239-2252.
[8]Ishimaru K, Hirotsu N, Madoka Y, et al. Loss of function of the IAA-glucose hydrolase gene TGW6 enhances rice grain weight and increases yield[J]. Nature Genetics, 2013, 45(6): 707-711.
[9]Wang Y, Xiong G, Hu J, et al. Copy number variation at the GL7 locus contributes to grain size diversity in rice[J]. Nature Genetics, 2015, 47(8): 944-948.
[10]Wang S, Li S, Liu Q, et al. The OsSPL16-GW7 regulatory module determines grain shape and simultaneously improves rice yield and grain quality[J]. Nature Genetics, 2015, 47(8): 949-954.
[11]Zhou Y, Miao J, Gu H, et al. Natural variations in SLG7 regulate grain shape in rice[J]. Genetics, 2015, 201(4): 1591-1599.
[12]Fan C, Xing Y, Mao H, et al. GS3, a major QTL for grain length and weight and minor QTL for grain width and thickness in rice, encodes a putative transmembrane protein[J]. Tag. Theoretical & Applied Genetics. Theoretische Und Angewandte Genetik, 2006, 112(6): 1164-1171.
[13]Hu Z, He H, Zhang S,et al. A kelch motif-containing serine/threonine protein phosphatase determines the large grain QTL trait in rice[J]. Journal of Integrative Plant Biology, 2012, 54(12): 979-990.
[14]Qi P, Lin Y S, Song X J, et al. The novel quantitative trait locus GL3.1 controls rice grain size and yield by regulating Cyclin-T1;3[J]. Cell Research, 2012, 22(12): 1666-1680.
[15]Zhang X, Wang J, Huang J, et al. Rare allele of OsPPKL1 associated with grain length causes extra-large grain and a significant yield increase in rice[J]. Proceedings of the National Academy of Sciences of the United States of America, 2012, 109(52): 21534-21539.
[16]Li Y, Fan C, Xing Y, et al. Natural variation in GS5 plays an important role in regulating grain size and yield in rice[J]. Nature Genetics, 2011, 43(12): 1266-1269.
[17]Xu C, Liu Y, Li Y, et al. Differential expression of GS5 regulates grain size in rice[J]. Journal of Experimental Botany, 2015, 66(9): 2611-2623.
[18]Sun L, Li X, Fu Y, et al. GS6, a member of the GRAS gene family, negatively regulates grain size in rice[J]. Journal of Integrative Plant Biology, 2013, 55(10): 938-949.
[19]Kesavan M,Song J T,Seo H S. Seed size: a priority trait in cereal crops[J]. Physiologia Plantarum, 2013, 147(2): 113-120.
[20]Li N and Li Y H. Signaling pathways of seed size control in plants[J]. Current Opinion in Plant Biology, 2016, 33: 23-32.
[21]裔传灯, 王德荣, 蒋伟, 等. 水稻粒形基因GS5的功能标记开发和单体型鉴定[J]. 中国水稻科学, 2016, 30(5): 487-492.
[22]裔传灯, 李玮, 王德荣, 等. 水稻粒形基因GS3的功能标记开发与鉴定[J]. 江苏农业科学,2016, 44(12): 64-67.
[23]裔传灯, 王德荣, 蒋伟, 等. 水稻粒形基因GW8的功能标记开发和单体型鉴定[J]. 作物学报, 2016, 42(9): 1291-1297.
[24]Ye S, Dhillon S, Ke X, et al. An efficient procedure for genotyping single nucleotide polymorphisms[J]. Nucleic Acids Research, 2001, 29(17): e88.
[25]卿冬进, 刘开强, 杨燕宇, 等. 基于PARMS技术的抗稻瘟病基因Pigm分子标记的开发[J]. 西南农业学报, 2018, 31(8): 1617-1621.
[26]Murray M G,Thompson W F. Rapid isolation of high molecular weight plant DNA[J]. Nucleic Acids Research, 1980, 8(19): 4321.
[27]李欣, 莫惠栋, 王安民, 等. 粳型杂种稻米品质性状的遗传表达[J]. 中国水稻科学, 1999, 13(4): 197-204.
[28]殷得所, 夏明元, 李进波, 等. 抗稻瘟病基因Pi9的STS连锁标记开发及在分子标记辅助育种中的应用[J]. 中国水稻科学, 2011, 25(1): 25-30.
[29]Huang K, Wang D, Duan P, et al. WIDE AND THICK GRAIN 1, which encodes an otubain-like protease with deubiquitination activity, influences grain size and shape in rice[J]. Plant Journal, 2017, 91(5): 849-860.