中国农业科学 ›› 2019, Vol. 52 ›› Issue (13): 2208-2219.doi: 10.3864/j.issn.0578-1752.2019.13.002

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

利用BSA法发掘野生大豆种子硬实性相关QTL

陈静静,刘谢香,于莉莉,卢一鹏,张嗣天,张昊辰,关荣霞(),邱丽娟   

  1. 中国农业科学院作物科学研究所/国家农作物基因资源与遗传改良重大科学工程/农业部种质资源利用重点实验室,北京 100081
  • 收稿日期:2019-02-28 接受日期:2019-04-14 出版日期:2019-07-01 发布日期:2019-07-11
  • 通讯作者: 关荣霞
  • 作者简介:陈静静,E-mail:1061184187@qq.com。
  • 基金资助:
    国家自然科学基金(31830066);中国农业科学院基本科研业务费(S2018QY03)

QTL Mapping of Hard Seededness in Wild Soybean Using BSA Method

CHEN JingJing,LIU XieXiang,YU LiLi,LU YiPeng,ZHANG SiTian,ZHANG HaoChen,GUAN RongXia(),QIU LiJuan   

  1. Institute of Crop Sciences, Chinese Academy of Agricultural Sciences/National Key Facility for Crop Gene Resources and Genetic Improvement (NFCRI)/Key Laboratory of Germplasm Utilization, Ministry of Agriculture, Beijing 100081
  • Received:2019-02-28 Accepted:2019-04-14 Online:2019-07-01 Published:2019-07-11
  • Contact: RongXia GUAN

摘要:

【目的】野生大豆的硬实性是大豆遗传改良利用中的重要限制因素。利用BSA法发掘与大豆种子硬实性相关的QTL,为野生大豆在大豆遗传改良中的合理利用奠定基础。【方法】利用栽培大豆中黄39与野生大豆NY27-38杂交构建F2和F7分离群体,从每个单株选取整齐一致的种子,取30粒种子置于铺有一层滤纸的培养皿中,加入30 mL蒸馏水,25℃培养箱中暗处理4 h,设3次重复,分别统计每个培养皿中正常吸胀和硬实种子数。在F2群体中,选取22个正常吸胀单株(吸胀率>90%)和16个硬实单株(吸胀率<10%);在F7群体中,选取20个完全吸胀单株(吸胀率=100%)和20个完全硬实单株(吸胀率=0%),单株DNA等量混合,分别构建2个吸胀和2个硬实DNA池。利用259对在亲本间有多态性的SSR标记对吸胀和硬实DNA池进行检测,筛选在吸胀和硬实DNA池间表现多态性的SSR标记;用192个SSR标记检测F7分离群体,构建遗传图谱,利用复合区间作图法定位大豆硬实相关QTL。【结果】利用F2个体构建的吸胀和硬实DNA池,在第2染色体16.3 Mb区间和第6染色体23.4 Mb区间分别检测到10个和8个在两池间有差异的SSR标记。利用这些标记检测F2群体,将第2染色体的QTL定位于Satt274与Sat_198间的276.0 kb区间,该区间包括已克隆的大豆硬实基因GmHs1-1,解释17.2%的表型变异。第6染色体的QTL位于标记BARCSOYSSR_06_0993与BARCSOYSSR_06_1068间,可解释17.8%的表型变异。利用F7株系构建的吸胀和硬实DNA池,在第2(27.4 Mb区间)、6(27.8 Mb 区间)和3染色体(18.2 Mb区间)分别检测到11个、9个和4个在两池间有多态性的SSR标记。利用F7群体构建包括192个SSR标记、覆盖2 390.2 cM的遗传图谱,共检测到3个硬实相关QTL,其中第2染色体定位到的QTL位于标记Satt274与Sat_198间,可解释23.3%的遗传变异。第6染色体定位到的QTL位于标记Sat_402与Satt557之间,可解释20.4%的表型变异。在第3染色体标记Sat_266与Sat_236间发现一个可以解释4.9%表型变异的QTL,与BSA法检测的结果相符。【结论】利用BSA法可以检测到传统遗传作图定位的所有与硬实性相关的QTL,证明BSA法发掘大豆种子硬实性主要QTL的高效性。

关键词: 大豆, 种子硬实, QTL定位

Abstract:

【Objective】 Hard seededness of wild soybean is an important effector that limits the utilization of wild resources in soybean genetic improvement. Bulked segregant analysis (BSA) was employed to identify major quantitative trait loci (QTLs) related with hard seededness in soybean, which laid a foundation for effective utilization of wild soybean germplasm in cultivated soybean improvement. 【Method】 F2 and F7 segregation populations were constructed from a cross between cultivated soybean Zhonghuang39 and wild soybean NY27-38. Uniformly sized seeds were selected from each line, and 30 seeds were soaked in a petri dish with 30 mL distilled water for 4 hours at 25℃. The assay was replicated 3 times. The number of permeable and impermeable seeds were counted. In F2 population, the first DNA pool was constructed from 22 individuals with permeable seeds (imbibition rate >90%), and second DNA pool was constructed from 16 individuals with impermeable seeds (imbibition rate <10%). In F7 population, 20 lines with permeable seeds (100% imbibition) and 20 lines with impermeable seeds (no imbibition) were used to construct two DNA pools, respectively. To detect genomic regions associated with hard seededness, these DNA bulks were genotyped with 259 polymorphic SSR markers to identify markers linked to QTL. A linkage map was constructed with 192 SSR markers, QTLs related with hard seededness were identified by composite interval mapping in F7 segregation population. 【Result】 Out of 259 SSR loci polymorphic between Zhonghuang39 and NY27-38, 10 and eight polymorphic SSR markers between the permeable and impermeable pools were detected in 16.3 Mb interval on chromosome 2 and 23.4 Mb interval on chromosome 6, respectively, in F2 population. The QTL region (276.0 kb) located between Satt274 and Sat_198 on chromosome 2 contained previously cloned gene GmHs1-1, the QTL explained 17.2% of the total genetic variation. The other QTL was mapped on chromosome 6 flanked by BARCSOYSSR_06_0993 and BARCSOYSSR_06_1068, accounting for 17.8% of the total genetic variation. In F7 population, eleven, nine and four SSR polymorphic markers between the permeable and impermeable pools were detected in 27.4 Mb interval on chromosome 2, 27.8 Mb interval on chromosome 6, 18.2 Mb interval on chromosome 3, respectively. A linkage map of 192 SSR markers and covering 2 390.2 cM was constructed through composite interval mapping in F7 population. Three QTLs related with hard seededness were detected. The QTL on chromosome 2 located between Satt274 and Sat_198, explained 23.3% of the total genetic variation; the QTL on chromosome 6 flanked by Sat_402 and Satt557, explained 20.4% of the total genetic variation; the QTL on chromosome 3 flanked by Sat_266 and Sat_236 accounted for 4.9% of the total genetic variation. 【Conclusion】 In this study, three QTLs related to soybean hard seededness were identified by both BSA and traditional linkage mapping, indicating that BSA is an effective strategy for identifying QTLs in soybean.

Key words: soybean, hard seededness, QTL mapping