利用高通量测序快速定位目标性状位点的研究

doi:10.7668/hbnxb.20191003

华北农学报 ›› 2019, Vol. 34 ›› Issue (6): 47-53. doi: 10.7668/hbnxb.20191003

• 作物遗传育种·种质资源·生物技术 • 上一篇下一篇

利用高通量测序快速定位目标性状位点的研究

林静^1,2, 赵青松¹, 张孟臣¹, 杨春燕¹, 赵团结²

1. 河北省农林科学院粮油作物研究所, 国家大豆改良中心石家庄分中心, 农业部黄淮海大豆生物学与遗传育种重点实验室, 河北省农作物遗传育种重点实验室, 河北石家庄 050035;
2. 南京农业大学国家大豆改良中心, 农业部大豆生物学与遗传育种重点实验室(综合), 国家作物遗传与种质资源重点实验室, 江苏南京 210095

收稿日期:2019-10-15 出版日期:2019-12-28
通讯作者: 杨春燕(1966-),女,黑龙江密山人,研究员,硕士生导师,主要从事大豆遗传育种研究;赵团结(1969-),男,安徽旌德人,教授,博士,博士生导师,主要从事大豆遗传育种研究。
作者简介:林静(1992-),女,福建龙岩人,博士,主要从事大豆遗传育种研究。
基金资助:
国家"863"计划项目（2012AA101106）；国家产业技术体系（CARS-004-PS06）；现代农业科技创新工程项目（2019-4-3-1）

Rapid Mining Genetic Locus for Controlling Target Traits Using Next-generation Sequencing (NGS)

LIN Jing^1,2, ZHAO Qingsong¹, ZHANG Mengchen¹, YANG Chunyan¹, ZHAO Tuanjie²

1. Institute of Cereal and Oil Crops, Hebei Academy of Agriculture and Forestry Sciences, National Center for Soybean Improvement, Shijiazhuang Sub-Center, Huang-huai-hai Key Laboratory of Biology and Genetic Improvement of Soybean, Ministry of Agriculture, The Key Laboratory of Crop Genetics and Breeding of Hebei, Shijiazhuang 050035, China;
2. National Center for Soybean Improvement, Key Laboratory of Biology and Genetics Improvement of Soybean, Ministry of Agriculture, National Key Laboratory for Crop Genetics and Germplasm, Nanjing Agricultural University, Nanjing 210095, China

Received:2019-10-15 Published:2019-12-28

摘要/Abstract

摘要： 为了快速鉴定目标性状遗传位点，开发与性状连锁的分子标记，用于剔除高世代选育株行中不利性状，从而加速育种进程。以大豆MS轮回群体中发生花色分离的F₆株行为研究材料，利用其衍生的F_6：7中20个紫花和17个白花纯合家系分别构建2个DNA混池，通过高通量重测序技术获得变异信息，明确SNP富集区，并在SNP富集区内开发分子标记对目标性状进行连锁分析。结果显示，在2个DNA混池间发现329 992个SNP位点，表明混池间的遗传背景已经非常相似，但未发现明显SNP富集区，表明可能存在较多假阳性位点；进一步对高质量（Quality> 100）的SNP变异位点进行筛选，并去除杂合SNP位点，最终获得3 371个可信位点。其中，位于13号染色上的SNP变异有700个（占比20.77%），并在20~30 Mb的物理区间形成一个最大的SNP富集区，推测调控花色的W1位点可能位于此区间内。利用该区间内SNP信息开发出dCAPS-1、dCAPS-2分子标记，连锁分析结果显示其与W1位点紧密连锁（W1-（0.4 cM）-dCAPS-1-（2.3 cM）-dCAPS-2），表明W1位点位于SNP富集区内。综上所述，通过构建高世代株行的分离群体混池，利用高通量测序方法可以快速定位控制目标性状的遗传位点，且开发的dCAPS标记可以有效剔除不利性状，从而加速遗传育种进程。

关键词: 高世代混池, 高通量测序, SNP变异, 遗传定位, 分子标记开发

Abstract: To rapidly determine the genetic loci for target traits and develop molecular markers linked to traits which could remove plants with disadvantage traits in high generation breeding line and accelerate the breeding process. In this study, a row of F₆ generation plants performing flower color segregation were selected from soybean MS recurrent population and individually harvested to form F_6:7 population. Among the F_6:7 population, 20 and 17 homozygous family lines with purple and white flower color, respectively, were selected to construct two DNA pools. Following, genetic variation were derived from next-generation sequencing and SNP enriched region were determined, furthermore, molecular markers were also developed to confirm the linkage between markers and target traits. High throughput sequencing analysis showed a total of 329 992 SNP variations were identified between the DNA pools of purple and white color, indicating that the genetic background between the DNA pools have a high degree of similarity. However, no significant SNP rich region was found possible due to there exist lots of false positive SNP. After screening SNP variations with high standard (Quality>100) and removing heterozygous SNP variations, a total of 3 371 confidence SNP variation were obtained. Among them, 700 (20.77%) distribute on chromosome 13, and most were clustered in the physical interval of 20 to 30 Mb, strongly indicating that W1 was localized in this region. Two new markers, dCAPS-1, dCAPS-2 in the region were developed and they were linked to W1 closely (W1 -(0.4 cM)-dCAPS-1-(2.3 cM)-dCAPS-2). In conclusion, the locus underlying the target traits could be rapidly determined through constructing two segregation pools from high generation line and genotyping by next-generation sequencing. Moreover, the disadvantage traits could be effective removed by the linked dCAPS markers which will accelerate breeding process.

Key words: Mixture pools of advance line, High-throughput sequencing, SNP variations, Genetic mapping, Molecular marker development

中图分类号:

Q78
S565.03

林静, 赵青松, 张孟臣, 杨春燕, 赵团结. 利用高通量测序快速定位目标性状位点的研究[J]. 华北农学报, 2019, 34(6): 47-53. doi: 10.7668/hbnxb.20191003.

LIN Jing, ZHAO Qingsong, ZHANG Mengchen, YANG Chunyan, ZHAO Tuanjie. Rapid Mining Genetic Locus for Controlling Target Traits Using Next-generation Sequencing (NGS)[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2019, 34(6): 47-53. doi: 10.7668/hbnxb.20191003.

http://www.hbnxb.net/CN/Y2019/V34/I6/47

参考文献

[1] 谷强平.中国大豆进口贸易影响因素及效应研究[D].沈阳:沈阳农业大学, 2015:10-30.
Gu Q P. A research on the influence factors and effect of China's soybean import trade[D]. Shenyang:Shenyang Agricultural University, 2015:10-30.
[2] 农业部.《全国种植业结构调整规划(2016-2020年)》[J].中国食品, 2016(10):149-154.
Ministry of Agriculture.《National planting structure adjustment plan (2016-2020)》[J].China Food,2016(10):149-154.
[3] Newton J. The footprint of Chinese demand for U.S. soybeans[J]. Population, 2015, 34(2):1171. doi:10.1103/PhysRevD.54.1379.
[4] Terzic D, Popovic V, Tatić M, Vasileva V, Rajicic V, Ugrenović V, Popović S, Avdić P. Soybean area yield and production in world[C]. Eco-Conference,2018.
[5] 孙其信.作物育种学[M].北京:高等教育出版社, 2011.
Sun Q X. Principles of crop breeding[M]. Beijing:Higher Education Press, 2011.
[6] Zhang H, Zhou P, Tu S H, Zheng J T, Zhang J F, Xie H A. Developing new restorer lines with blast-resistance gene Pi9 for hybrid rice by marker assistance selection (MAS)[J]. Molecular Plant Breeding, 2015,13(9):1918-1922.
[7] 陈晖,陈美霞,陶爱芬,张广庆,徐建堂,祁建民,方平平.长果种黄麻SRAP标记遗传连锁图谱的构建及3个质量性状基因定位[J].中国农业科学, 2011,44(12):2422-2430.
Chen H, Chen M X, Tao A F, Zhang G Q, Xu J T, Qi J M, Fang P P. Construction of a molecular linkage map using SRAP and mapping of three qualitative traits in Corchorus olitoriusL.[J]. Scientia Agricultura Sinica, 2011,44(12):2422-2430.
[8] 杨光华,范荣,杨小锋,侯军亮,袁士臣,曹明,王学林,李劲松.甜瓜果实颜色3个质量性状基因的定位[J].园艺学报, 2014,41(5):898-906.
Yang G H, Fan R, Yang X F, Hou J L, Yuan S C, Cao M, Wang X L, Li J S. Construction of a highly dense genetic map using SNP and mapping of three qualitative traits in Cucumis melo[J]. Acta Horticulturae Sinica, 2014,41(5):898-906.
[9] 倪西源,王学德,程超华,王晓玲,张昭伟.棉花分子标记图谱的构建和一些重要性状的定位[J].棉花学报, 2007,19(1):71-73. doi:10.3969/j.issn.1002-7807.2007.01.015.
Ni X Y, Wang X D, Cheng C H, Wang X L, Zhang S W. Constructing of DNA molecular marker linkage map and mapping of qualitative and quantitative traits in upland cotton[J]. Cotton Science, 2007,19(1):71-73.
[10] Quarrie S, Quarrie S P, Radosevic R, Rancic D, Kaminska A, Barnes J D, Leverington M,Ceoloni C, Dodig D. Dissecting a wheat QTL for yield present in a range of environments:from the QTL to candidate genes[J]. J Exp Bot, 2006, 57(11):2627-2637. doi:10.1093/jxb/erl026.
[11] Yin Z G, Qi H D, Chen Q S, Zhang Z G, Jing H W, Zhu R S, Hu Z B, Wu X X, Li C D, Zhang Y, Liu C Y, Hu G H, Xin D W, Qi Z M. Soybean plant height QTL mapping and meta-analysis for mining candidate genes[J]. Plant Breeding, 2017, 136(5):688-698. doi:10.1111/pbr.12500.
[12] 陈广凤,田纪春.基于SNP标记小麦自然群体遗传多样性及复合图谱的构建[J].分子植物育种, 2015,13(7):1441-1449. doi:10.13271/j.mpb.013.001441.
Chen G F, Tian J C. Genetic analysis of natural population of wheat and construction of composite map using SNP Markers[J]. Molecular Plant Breeding, 2015,13(7):1441-1449.
[13] 李文明.玉米CMS-S育性恢复基因定位分析及EMS突变体筛选[D].武汉:华中农业大学, 2018.
Li W M. Genetic mapping and identification of EMS mutants for fertility restoration of CMS-S in maize[D]. Wuhan:Huazhong Agricultural University, 2018.
[14] Li J M, Thomson M, Mccouch S. Fine mapping of a grain-weight quantitative trait locus in the pericentromeric region of rice Chromosome 3[J]. Genetics, 2004, 168(4):2187-2195. doi:10.1534/genetics.104.034165.
[15] 杨梅.玉米株高QTL qPH3.2和qPH3.3的精细定位[D].武汉:华中农业大学, 2017.
Yang M. Fine mapping of the major QTLqPH3.2 and qPH3.3 for plant height in maize (Zea mays L.)[D]. Wuhan:Huazhong Agricultural University, 2017.
[16] 刘立科,侯宁,刘建成,刘根齐,刘春光.小麦D2型细胞质雄性不育恢复基因近等基因系筛选和遗传背景的分子检测[J].麦类作物学报, 2006, 26(5):1-4,15. doi:10.3969/j.issn.1009-1041.2006.05.001.
Liu L K, Hou N, Liu J C, Liu G Q, Liu C G. Screening of the near-isogenic lines (NILs) of fertility restorer genes for D2-type cytoplasmic male sterility (CMS) in wheat and molecular characterization of the genetic backgrounds[J]. Journal of Triticeae Crops, 2006, 26(5):1-4,15.
[17] Yamanaka N, Watanabe S, Toda K, Hayashi M, Fuchiqami H, Takahashi R, Harada K. Fine mapping of the FT1 locus for soybean flowering time using a residual heterozygous line derived from a recombinant inbred line[J]. Theoretical and Applied Genetics, 2005, 110(4):634-639. doi:10.1007/s00122-004-1886-3.
[18] Li H, Durbin R. Fast and accurate short read alignment with Burrows-Wheeler transform[J]. Bioinformatics, 2009, 25(14):1754-1760. doi:10.1093/bioinformatics/btp324.
[19] Voorrips R E. MapChart:software for the graphical presentation of linkage maps and QTLs[J]. Journal of Heredity 2002, 93(1):77-78. doi:10.1093/jhered/93.1.77.
[20] 史宏,任小俊,马俊奎,王勇,赵晶云,刘学义.大豆重组自交系Jinf F₁₀抗大豆孢囊线虫4号生理小种抗性的分析研究[J].华北农学报, 2008, 23(3):176-180. doi:10.7668/hbnxb.2008.03.042.
Shi H, Ren X J, Ma J K, Wang Y, Zhao J Y, Liu X Y. Studies on relationship between agronomic traits and resistance of SCN4 race in soybean canopies[J]. Acta Agriculturae Boreali-Sinica, 2008, 23(3):176-180.
[21] Nguyen T T T, Koizumi S, La T N, Zenbayashi K S, Ashizawa T, Yasuda M, Imazaki I, Miyasaka A. Pi35(t), a new gene conferring partial resistance to leaf blast in the rice cultivar Hokkai 188[J]. Theoretical and Applied Genetics, 2006, 113(4):697-704. doi:10.1007/s00122-006-0337-8.
[22] Luan J W, Wang F, Li Y J, Zhang B, Zhang J R. Mapping quantitative trait loci conferring resistance to Rice black-streaked virus in maize (Zea mays L.)[J]. Theoretical and Applied Genetics, 2012, 125(4):781-791.
[23] Song J, Liu Z X, Hong H L, Ma Y S, Tian L, Li X X, Li Y H, Guan R X, Guo Y, Qiu L J. Identification and validation of loci governing seed coat color by combining association mapping and bulk segregation analysis in soybean[J]. PLoS One, 2016, 11(7):e0159064.
[24] Sundaramoorthy J, Park G T, Lee G D, Kim J H, Seo H S, Song J T. Genetic and molecular regulation of flower pigmentation in soybean[J]. Journal of the Korean Society for Applied Biological Chemistry, 2015, 58(4):555-562. doi:10.1007/s13765-015-0077-z.
[25] Schmutz J, Cannon S B, Schlueter J, Ma J X, Mitros T, Nelson W, Hyten D L, Song Q J, Thelen J J, Cheng J L, Xu D, Hellsten U, May G D, Yu Y, Sakurai T, Umezawa T, Bhattacharyya M K, Sandhu D, Valliyodan B, Lindquist E, Peto M, Grant D, Shu S Q, Goodstein D, Barry K, Futrell-Griggs M, Abernathy B, Du J C, Tian Z X, Zhu L C, Gill N, Joshi T, Libault M, Sethuraman A, Zhang X C, Shinozaki K, Nguyen H T, Wing R A, Cregan P, Specht J, Grimwood J, Rokhsar D, Stacey G, Shoemaker R C, Jackson S A. Genome sequence of the palaeopolyploid soybean[J]. Nature,2010, 463(7278):178-183. doi:10.1038/nature08957.
[26] 吴亚君,杨艳歌,李莉,王斌,刘鸣畅,陈颖.高通量二代测序基因条码技术在油料作物种类鉴别中的应用[J].食品科学, 2014, 35(24):348-352. doi:10.7506/spkx1002-6630-201424067.
Wu Y J, Yang Y G, Li L, Wang B, Liu M C, Chen Y. Application of high throughput next-generation sequencing based on DNA barcoding technology in species identification of edible oils[J]. Food Science, 2014, 35(24):348-352.
[27] Wang L, Cheng Y B, Ma Q B, Mu Y H, Huang Z F, Xia Q J, Zhang G Y, Nian H. QTL fine-mapping of soybean (Glycine max L.) leaf type associated traits in two RILs populations[J]. BMC Genomics, 2019, 20:260. doi:10.1186/s12864-019-5610-8.

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