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作物学报 ›› 2023, Vol. 49 ›› Issue (5): 1222-1230.doi: 10.3724/SP.J.1006.2023.24140

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

花生InDel标记开发及其在含油量QTL定位中的应用

陶顺玉(), 吴贝, 刘念, 罗怀勇, 黄莉, 周小静, 陈伟刚, 郭建斌, 喻博伦, 雷永, 廖伯寿, 姜慧芳()   

  1. 中国农业科学院油料作物研究所/农业农村部遗传育种重点实验室, 湖北武汉 430062
  • 收稿日期:2022-06-10 接受日期:2022-07-21 出版日期:2023-05-12 网络出版日期:2022-09-23
  • 通讯作者: *姜慧芳, E-mail: peanutlab@oilcrops.cn
  • 作者简介:E-mail: ynautsy@163.com
  • 基金资助:
    广东省重点领域研发计划项目(2020B020219003);国家自然科学基金项目(31801403);国家自然科学基金项目(31871666);农作物种质资源保护项目(2017NWB033);国家农作物种质资源共享服务平台(NICGR2017-36);财政部和农业农村部国家现代农业产业技术体系建设专项(CARS-13-花生种质资源评价);中国农业科学院创新工程项目(2022-2060299-089-031)

Development and employment of InDel marker in peanut QTL mapping of oil content

TAO Shun-Yu(), WU Bei, LIU Nian, LUO Huai-Yong, HUANG Li, ZHOU Xiao-Jing, CHEN Wei-Gang, GUO Jian-Bin, YU Bo-Lun, LEI Yong, LIAO Bo-Shou, JIANG Hui-Fang()   

  1. Oil Crops Research Institute, Chinese Academy of Agricultural Sciences/Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture and Rural Affairs, Wuhan 430062, Hubei, China
  • Received:2022-06-10 Accepted:2022-07-21 Published:2023-05-12 Published online:2022-09-23
  • Contact: *E-mail: peanutlab@oilcrops.cn
  • Supported by:
    Key Area Research and Development Program of Guangdong Province(2020B020219003);National Natural Science Foundation of China(31801403);National Natural Science Foundation of China(31871666);Crop Germplasm Resources Protection Project(2017NWB033);Plant Germplasm Resources Sharing Platform(NICGR2017-36);China Agriculture Research System of MOF and MARA(CARS13);Innovation Project of Chinese Academy of Agricultural Sciences(2022-2060299-089-031)

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摘要：

培育高含油量品种是花生重要的育种目标, 开发稳定高效的分子标记对于标记辅助选择花生高含油量品种具有重要价值。本研究针对2个含油量差异显著的亲本徐花13 (高含油量)和中花6号(低含油量), 利用PacBio三代测序技术检测基因组结构变异, 分别从徐花13和中花6号中检测到35,794个和74,703个结构变异。根据双亲结构变异信息, 针对前期定位的含油量区间开发InDel (插入/缺失)标记84个, 其中9个InDel标记的扩增片段证实在双亲间具有多态性。基于RIL (重组自交系)群体中的杂合残余获得的NIL (近等基因系)构建的一个包含1160个单株的F2群体, 使用9个多态性InDel标记鉴定其基因型, 构建了总长度为149.84 cM的局部遗传连锁图谱。结合F2群体的含油量表型数据, 将该含油量QTL定位在标记M23至M11之间, 位于A08染色体1.2 Mb区段内。本试验证实了InDel标记开展花生含油量QTL定位的可行性, 其定位的含油量位点及其紧密连锁的InDel标记能够为花生分子标记辅助育种提供理论和技术指导。

关键词: 花生, 基因组测序, InDel, 含油量

Abstract:

The developing of stable and efficient molecular markers is of great value for marker-assisted selection of peanut varieties with high oil content. In this study, two parents with significant differences in oil content, Xuhua 13 (high oil content) and Zhonghua 6 (low oil content), were used to detect genomic structural variation by PacBio third-generation sequencing technology, and 35,794 and 74,703 structural variations were detected in Xuhua 13 and Zhonghua 6, respectively. 84 InDel markers in the target region were detected based on the parents’ structural variations and the previous QTL mapping information. And 9 InDel markers were found to be polymorphic between the parents. Meanwhile, F2 population with 1160 individual plants was constructed based on near isogenic lines (NILs) derived from heterozygous residuals in the recombined inbred line (RIL) population. The polymorphic markers were used to genotype the population and to construct the genetic linkage map with 149.84 cM. Combining with phenotype data of population for oil content, QTL (qOCA08) was fine mapped between marker M23 and marker M11, which was located on 1.2 Mb interval of chromosome A08. This study demonstrated the feasibility of InDel marker for peanut QTL mapping. The locus of oil content and closely linked InDel markers can provide the theoretical and technical guidance for peanut molecular marker-assisted breeding.

Key words: peanut, genome resequencing, InDel, oil content

表1

亲本测序信息"

样品
Sample name		 碱基数目
Base number		 Reads数
Clean reads		 比对率
Mapping rate (%)		 覆盖度
Coverage
(%)		 结构变异
Structural variation		 插入
Insertion		 缺失
Deletion
徐花13 Xuhua 13 81,826,464,736 5,615,798 98.03 96.91 35,794 12,293 4496
中花6号 Zhonghua 6 68,494,830,848 4,225,186 97.58 96.59 74,703 26,833 12,906

图1

亲本中插入、缺失长度分布 A: 徐花13插入变异的长度分布; B: 徐花13缺失变异的长度分布; C: 中花6号插入变异的长度分布; D: 中花6号缺失变异的长度分布。"

图2

结构变异发生在基因组不同区域的比例 A: 徐花13的结构变异在基因组不同区域的比例; B: 中花6号的结构变异在基因组不同区域的比例。"

图3

亲本中9个多态性InDel标记PCR扩增结果"

表2

亲本中具有多态性的InDel标记信息"

引物名称
Primer name		 引物序列
Primer sequences (5'-3')		 引物位置(A08)
Physical position (A08) (bp)		 差异大小
Difference in size (bp)
M9-F AGTATTGAAGCGGCAGGAGG 37,791,529 18
M9-R TTGCACTTCGATGTGGAGGAT
M21-F TAGTTCTCTCCGGATGCCTCA 42,578,036 26
M21-R TGCCATGAAATTACTCATTACTCTT
M6-F CAGAGAGACAGAGAGCGAGC 44,213,248 466
M6-R TGAGCCGGATGAGTAAGACA
M22-F AATAAGTTTGTTGGGAGAATGAAAT 44,339,712 24
M22-R GCTCCCATTGTCTAGCGAAAAC
M23-F TGCTGTGTGTAATAGTTAGAGGCT 44,988,232 28
M23-R TGGGGTGAGTGAATTATACGTGA
M11-F TGTGAGTACCGACATACACTCA 46,212,241 154
M11-R AATTTGATTATTGGACATCCCCT
M13-F GTTGAGACCGGAACACAGGA 46,400,655 362
M13-R CTATCCACTAGGTTCTCCAGCA
M14-F TGTCAATGTGTTATCATCTCAATTT 46,555,343 2353
M14-R TAATTGGCACCACGCAATGTC
M8-F CCAAGTCCTTTGCAGCTCTC 49,660,135 15
M8-R AAATTACTTTCAGTAAATGTCTAGC

图4

F2群体含油量表型的频率分布"

图5

F2群体定位结果"

图6

M11位点等位基因型含油量的表型效应 **代表在0.01概率水平著差异(t检验)。"

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