作物分子身份证构建软件ID analysis的编制

doi:10.3864/j.issn.0578-1752.2016.12.002

摘要/Abstract

摘要： 【目的】以作物品种或资源的SSR分子标记为基础数据，应用Visual Basic6.0开发作物分子身份证构建配套软件ID analysis，快速准确地筛选引物组合并高效鉴定品种。【方法】作物分子身份证理论由陈庆山提出，其原理是利用SSR标记在材料中的多态性特点，将多个标记进行排列组合，快速有效地区分品种资源的算法——逐步扩增法。通过对40份黑龙江省大豆品种的40对引物数据进行分子身份证构建，阐述该软件的详细操作过程。【结果】利用Visual Basic6.0编程软件设计人机交互界面，通过编程实现核心算法，开发ID analysis软件，软件集成全库构建、部分构建、ID判定、数据合并等功能。其中，全库构建是软件的核心功能，通过全库构建可以快速分析区分材料的最少标记；部分构建可以选择部分目标材料进行区分；ID判定可以在分子身份证数据库构建完成的前提下，对未知材料进行SSR分析，根据获得的分子身份证数据来确定材料是哪个品种或近似品种；数据合并功能可以将多次试验数据整合成一个数据集。利用软件全库构建功能对范例数据进行分析得到以下结果：①在40对引物对40个大豆品种的分子身份证构建中，共有13个引物由于缺失过多，不符合标准被剔除，剔除引物为：Sat_111、Sat_218、Satt231、Satt685、Satt514、Satt551、Satt077、Satt358、Satt424、Satt100、Satt838、Satt893和Satt891。②共有6个引物由于与其他引物相似系数过高，不符合标准被剔除，剔除引物为：Satt253、Satt192、Satt417、Sat_229、Satt127和Satt496。③在分析的40个品种中，共有5个品种具有7个特异等位基因，分别为引物Satt516在材料东农36中显示特异条带3；引物Satt253在材料东农36中显示特异条带1；引物Sat_229在材料嫩丰17中显示特异条带1；引物Satt192在材料东农42中显示特异条带3；引物Satt206在材料北丰19中显示特异条带1；引物Satt244在材料北丰19中显示特异条带4；引物Satt363在材料黑河14中显示特异条带1，因此，可以通过这些特异等位基因直接确定需要鉴定的品种。④仅需7对引物便可将40份大豆品种完全区分开，引物组合为：Satt398、Satt380、Satt453、Satt288、Satt244、Sat_092和Satt206。【结论】编制了用于作物分子身份证构建的软件ID analysis，软件界面友好、使用简便、高效、灵活。实现单个软件完成作物分子身份证的构建，达到了对资源品种鉴定的目的。随着技术的发展和毛细管电泳的应用，开发全自动分子身份证分析系统甚至开发基于分子身份证理论的快速资源鉴定仪将成为可能。

关键词: 大豆, 作物, SSR标记, 分子身份证, 软件开发

Abstract: 【Objective】Based on the SSR molecular marker data of crops resources, software ID analysis was developed using visual Basic6.0 for crop molecular identity construction, which could screen primer combinations rapidly and accurately for efficient cultivar identification. 【Method】The crop molecular ID theory was proposed by Mr. Qingshan Chen. SSR markers in crop varieties showed high polymorphism characteristics, and a set of markers was permutated and combined to quickly and effectively divide varieties with step amplification method. Finally, 40 paires of SSR data of 40 soybean varieties in Heilongjiang province were used for identity construction with the software. 【Result】 ID analysis software was developed with core algorithm by using Visual Basic 6.0 to design man-machine interactive interface. This software has integrated full-library construction, partial-library construction, molecular ID determination, and database merging function. The full-library construction was the core functions together which could quickly obtained the minimum SSR prime combination to distinguish all the varieties. Partial-library construction could be used for some target varieties identification. For an unknown materials, with the already existing molecular identity database and the SSR analysis data, molecular ID determination could be used to determine the variety names or similar varieties. Database merging could be used to integrate several experimental data into a data set. The following results were analyzed by full-library construction with case data. First, among the 40 pairs of SSR data of 40 soybean cultivars, a total of 13 primers were excluded because missing data were too much and did not meet the standards, they were Sat_111, Satt218, Satt231, Satt685, Satt514, Satt551, Satt077, Satt358, Satt424, Satt100, Satt838 ,Satt893, and Satt891. Second, 6 primers were excluded because they showed high similarity coefficient with other primers, and they were Satt253, Satt192, Satt417, Sat_229, Satt127, and Satt496. Third, 5 varieties showed 7 specific alleles among all 40 varieties. They were, allele 3 of Satt516 and allele 1 of Satt253 showed in Dongnong36, allele 1 of Sat_229 showed in Nenfeng 17, allele 3 of Satt192 showed in Dongnong42, allele 1 of Satt206 and allele 4 of Satt244 showed in Beifeng 19, and allele 1 of Satt363 showed in Heihe14. So these specific allelic genes could directly identify the varieties. Forth, only seven pairs of SSR primers could distinguish 40 soybean varieties completely. The primer combinations were Satt398, Satt380, Satt453, Satt288, Satt244, Sat_092, and Satt206. 【Conclusion】 In this research, the software analysis ID was developed to construct crop molecular identity. The software has a friendly interface and easy to be used, high efficiency and flexible. The construction of the crop molecular identity can be realized completely by using a single software, and thus achieving the purpose of variety identification. With the development of technology of the application of capillary electrophoresis and the molecular identity theory, the development of an automatic molecular identity analysis system and even a rapid resource identification system will become possible.

Key words: soybean, crops, SSR markers, molecular identity, software development

胡振帮，高运来，齐照明，蒋洪蔚，刘春燕，辛大伟，胡国华，潘校成，陈庆山. 作物分子身份证构建软件ID analysis的编制[J]. 中国农业科学, 2016, 49(12): 2255-2266.

HU Zhen-bang, GAO Yun-lai, QI Zhao-ming, JIANG Hong-wei, LIU Chun-yan, XIN Da-wei, HU Guo-hua, PAN Xiao-cheng, CHEN Qing-shan. Software Development of -ID Analysis for Crop Molecular Identity Construction[J]. Scientia Agricultura Sinica, 2016, 49(12): 2255-2266.

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