中国农业科学 ›› 2021, Vol. 54 ›› Issue (22): 4880-4893.doi: 10.3864/j.issn.0578-1752.2021.22.014

• 园艺 • 上一篇    下一篇

葡萄无核基因分子标记的通用性鉴定

陈豆豆(),关利平,贺亮亮,宋银花,章鹏,刘三军()   

  1. 中国农业科学院郑州果树研究所,郑州 450009
  • 收稿日期:2021-01-25 接受日期:2021-04-08 出版日期:2021-11-16 发布日期:2021-11-19
  • 通讯作者: 刘三军
  • 作者简介:陈豆豆,E-mail: chendoudou21@163.com
  • 基金资助:
    河南省科技攻关项目(202102110196);国家现代农业产业技术体系(CARS-30-17);中国农业科学院创新工程“葡萄资源及育种”(CAAS-ASTIP-2020-ZFRI)

Commonality Identification of Molecular Markers Linked to Seedless Genes in Grape

CHEN DouDou(),GUAN LiPing,HE LiangLiang,SONG YinHua,ZHANG Peng,LIU SanJun()   

  1. Zhengzhou Fruit Research Institute, Chinese Academy of Agricultural Sciences, Zhengzhou 450009
  • Received:2021-01-25 Accepted:2021-04-08 Online:2021-11-16 Published:2021-11-19
  • Contact: SanJun LIU

摘要:

【目的】无核是鲜食葡萄的重要农艺性状,无核基因分子标记的准确性将影响无核葡萄早期选择的效果。本研究对前人开发的5个葡萄无核基因分子标记在183份葡萄材料中进行通用性鉴定,旨在明确各个分子标记的适用范围,为利用分子标记辅助无核葡萄育种提供参考依据。【方法】以96个葡萄品种(其中无核63个,有核33个)及‘红地球’ב黎明无核’87个F1单株(其中无核61株,有核26株)的叶片DNA为模板,利用已报道的5个葡萄无核基因分子标记(SCF27-2000、GSLP1-569、VMC7f2、p3_VvAGL11和5U_VviAGL11)的引物进行PCR扩增,采用1.5%琼脂糖凝胶电泳、毛细管电泳和8%聚丙烯酰胺凝胶电泳技术检测PCR产物,分析各个样品的无核特异条带,鉴定5个分子标记的准确率。【结果】SCAR标记GSLP1-569对96个葡萄品种的检测准确率为40.6%,无核检测率为63.6%。SCAR标记SCF27-2000对96个葡萄品种及87个F1杂交单株的检测准确率分别为71.9%和76.54%,无核检测率分别为70.5%和78.5%。SSR标记VMC7f2在96个葡萄品种中鉴定出8个等位基因,经卡方检验,189-bp等位基因与葡萄无核性状显著关联,其检测准确率为85.4%,无核检测率为85.5%。SSR标记p3_VvAGL11在96个葡萄品种中鉴定出7个等位基因,经卡方检验,187-bp等位基因与葡萄无核性状显著关联,其检测准确率为89.6%,无核检测率为90.7%,在F1杂交单株中其鉴定准确率为87.65%,无核检测率为91.1%,假阳性率为6.17%,假阴性率为6.17%。SSR标记5U_VviAGL11在96个葡萄品种中鉴定出17个等位基因,经卡方检验,306-bp等位基因与葡萄无核性状显著关联,其鉴定准确率为88.5%,无核检测率为90.6%,在F1杂交单株中检测准确率为88.89%,无核检测率为92.7%,假阳性率为4.94%,假阴性率为6.17%。【结论】SSR标记p3_VvAGL11和5U_VviAGL11在葡萄种质及遗传群体中无核分型的准确性及无核检测的效率较高,适用范围较广,在无核基因分子标记辅助选择中可以考虑优先使用。

关键词: 葡萄, 无核性, 分子标记, 通用性, 鉴定

Abstract:

【Objective】Seedlessness is an important agronomic trait in table grape breeding studies. The accuracy of molecular markers linked to seedless genes will substantially influence the early selection of seedless grapes. In this study, five previously reported seedless molecular markers against 183 genotypes were assessed to determine their efficacy for the selection of seedless plants and to provide alternative strategies for the breeding of seedless grapevines based on marker-assisted selection. 【Method】 Leaf samples for DNA extraction were obtained from 96 grape cultivars (63 seedless and 33 seeded cultivars) and 87 F1 hybrids (61 seedless and 26 seeded hybrids) from a cross between ‘Red Globe’ and ‘Dawn Seedless’. The previously reported primer pairs for five markers (SCF27-2000, GSLP1-569, VMC7f2, p3_VvAGL11, and 5U_VviAGL11) were used for PCR amplification. The presence of markers in cultivars and hybrid genotypes was detected by running the PCR products on 1.5% agarose gels, 8% polyacrylamide gels, or by fluorescence capillary electrophoresis. 【Result】 The primer for the SCAR marker GSLP1-569 amplified a 569-bp segment in the 96 grape cultivars, which showed 40.6% accuracy and 63.6% efficiency in the detection of seedless cultivars. The primers for the SCAR marker SCF27-2000 amplified a 2000-bp segment in the 96 grape cultivars and 87 F1 hybrids, with an accuracy of 71.9% and an efficiency of 76.54% for detection of the seedless trait in the 96 cultivars, and an accuracy of 70.5% and efficiency of 78.5% in the 87 F1 hybrids. Among the 96 assessed cultivars, the SSR marker VMC7f2 identified eight alleles, a 189-bp allele of which was found to be most significantly associated with seedlessness based on a Chi-square (χ2) independence test, and detected seedless genotypes with an accuracy of 85.4% and an efficiency of 85.5%. Among the 96 cultivars, the SSR marker p3_VvAGL11 identified seven alleles, among which, a 187-bp allele was found to be most significantly associated with seedlessness and detected seedless genotypes with an accuracy of 89.6% and efficiency of 90.7%. Among the F1 hybrids, the accuracy and efficiency of this marker were 87.65% and 91.1%, respectively, whereas it showed false-positive and false-negative rates of 6.17% and 6.17%, respectively. Among the 96 grape cultivars, the SSR marker 5U_VviAGL11 identified 17 alleles, of which, a 306-bp allele was found to be most significantly associated with the detection of seedlessness, identifying seedless genotypes with an accuracy of 88.5% and an efficiency of 90.6%. Among the F1 hybrids, the accuracy and efficiency of 5U_VviAGL11 were 88.89% and 92.7%, respectively, with false-positive and false-negative rates of 4.94% and 6.17%, respectively. 【Conclusion】 The SSR markers 5U_VviAGL11 and p3_VvAGL11 showed higher accuracy and efficiency with respect to the detection of grape germplasm and genetic population. These markers should be given priority for future molecular marker-assisted selection based on seedless genes in the grapevine.

Key words: Vitis vinifera L., seedlessness, molecular marker, commonality, identification