新陆早棉花品种DNA指纹图谱的构建及遗传多样性分析

引用本文

聂新辉, 尤春源, 李晓方, 秦江鸿, 黄聪, 郭欢乐, 王夏青, 赵文霞, 林忠旭. 新陆早棉花品种DNA指纹图谱的构建及遗传多样性分析. 作物学报, 40(12): 2104-2117
NIE Xin-Hui, YOU Chun-Yuan, LI Xiao-Fang, QIN Jiang-Hong, HUANG Cong, GUO Huan-Le, WANG Xia-Qing, ZHAO Wen-Xia, LIN Zhong-Xu. Construction of DNA Fingerprinting and Analysis of Genetic Diversity for Xinluzao Cotton Varieties. Acta Agronomica Sinica, 40(12): 2104-2117 复制到剪切板

Permissions

作物学报编辑部

新陆早棉花品种DNA指纹图谱的构建及遗传多样性分析

聂新辉^1,^2,^*, 尤春源^2,^*, 李晓方³, 秦江鸿², 黄聪¹, 郭欢乐¹, 王夏青¹, 赵文霞¹, 林忠旭^1,^*

¹华中农业大学作物遗传改良国家重点实验室, 湖北武汉 430070

²石河子农业科学研究院棉花研究所, 新疆石河子 832000

³长江大学, 湖北荆州 434025

^*通讯作者(Corresponding author): 林忠旭, E-mail:linzhongxu@mail.hzau.edu.cn^** 同等贡献(Contributed equally to this work)

收稿日期:2014-09-16

基金:本研究由国家公益性行业(农业)科研专项(201303008), “十二五”兵团重点领域科技攻关项目(2011BA001)和新疆兵团石河子科技攻关项目(2009NY05)资助;

摘要

以新疆2013年前审定的51个新陆早常规棉花品种为材料, 从5000对SSR引物中筛选出多态性高、稳定性好、且定位在棉花26条染色体上(每条染色体上选择2~3对)的75对核心引物, 检测到多态性位点226个, 每个标记检测到的基因型位点数在2~12之间, 平均为3.01个; 引物多态信息量(PIC)值介于0.0799~0.8752之间, 平均值为0.6624。结果显示, 在51份新陆早棉花常规品种中, 可利用特征引物将21份品种一次性区分开。利用40对引物可以完全区分开新陆早51份常规品种, 并构建供试品种的指纹图谱。同时利用NTSYS-pcV2.10软件聚类分析表明, 51个新陆早棉花品种遗传相似系数变化范围为0.4269~0.9873, 平均值为0.7071, 说明新陆早棉花品种之间遗传多样性较狭窄; 遗传相似系数矩阵和聚类分析将51个新陆早品种分为4大类型, 与原品种选育系谱高度吻合。

关键词: 新陆早常规品种; SSR; DNA指纹图谱; 遗传多样性

Construction of DNA Fingerprinting and Analysis of Genetic Diversity for Xinluzao Cotton Varieties

NIE Xin-Hui^1,^2,^**, YOU Chun-Yuan^2,^**, LI Xiao-Fang³, QIN Jiang-Hong², HUANG Cong¹, GUO Huan-Le¹, WANG Xia-Qing¹, ZHAO Wen-Xia¹, LIN Zhong-Xu^1,^*

¹National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan 430070, China

²Cotton Research Institute, Shihezi Academy of Agricultural Sciences, Shihezi 832000, China

³Yangtze University, Jingzhou 434025, China

Abstract

Fifty-one Xinluzao conventional cotton varieties authorized before 2013 were detected with 75 pairs of primers with high polymorphism, good repeatability, and even distribution on 26 chromosomes (2-3 pairs from each chromosome) selected from 5000 pairs of SSR primers. A total of 226 polymorphic loci were obtained, and each marker detected 2-12 polymorphic loci with an average of 3.01; the polymorphism information content ranged from 0.0799 to 0.8752, with an average of 0.6624. The results showed that 21 varieties were differentiated by characteristic primers, and the rest 30 varieties could be identified by primer conbinations. In the end, 40 specific primers and combinated primers could completely differentiate the 51 Xinluzao cotton varieties from each other. DNA fingerprinting of the 51 Xinluzao conventional varieties were constructed with the 40 pairs of SSR markers. NTSYS-pcV2.10 software was used to analyze genetic diversity of the 51 conventional varieties, the results indicated that genetic similarity coefficient for the varieties ranged from 0.4269 to 0.9873, with an average of 0.7071, showing the narrow genetic diversity in Xinluzao cotton varieties. The 51 Xinluzao cotton varieties were divided into four types with the genetic similarity coefficient matrix and cluster analysis, which were strongly consistent with their pedigrees.

Keyword: Xinluzao conventional cotton varieties; SSR; DNA fingerprinting; Genetic diversity

Show Figures

棉花是我国重要的经济作物, 优质、高产棉花品种的选育在棉花产业可持续稳定发展中发挥重要作用。随着棉花产业形势的发展, 新品种审定速度快、数量多, 而骨干亲本数量有限且反复利用, 导致品种间遗传差异变小; 随着转基因技术在棉花育种中的应用, 在原品种基础上改良少数或单个性状的衍生品种增多, 完全依据形态性状进行棉花品种的辨别越来越困难^{[ 1]}。同时, 推向生产的新品种更新换代快、数量多, 导致种子市场“混、杂、乱”局面, 阻碍棉花产业的健康发展。传统的棉花品种真实性和纯度是通过田间种植的方法鉴定的, 耗时长、成本高、时效性差, 易受环境因素的影响^{[ 2]}。

近年来, 随着分子生物学的不断发展, 分子标记技术因不受环境条件的影响且多态性标记可反映DNA水平上生物个体间的遗传差异而被广泛应用, 其中SSR标记被认为是品种鉴定较为理想的标记之一^{[ 3]}。目前, 分子标记技术广泛应用在棉花品种指纹图谱构建和遗传多样性分析中, 例如, 郭旺珍等^{[ 4]}最早在1996年利用RAPD技术对中国9个棉花主栽品种的指纹图谱进行了研究。Multani等^{[ 5]}利用RAPD标记构建了澳大利亚13个陆地棉品种和1个海岛棉品种的指纹图谱。Iqbal等^{[ 6]}对22个陆地棉品种和1个亚洲棉品种进行RAPD分析, 认为陆地棉品种的遗传基础很窄。武耀廷等^{[ 7]}利用SSR标记检测了30个陆地棉栽培品种和4个杂交种亲本的多态性, 并建立了杂交种的指纹图谱, 用于它们的分子鉴定和纯度检测。马轩等^{[ 8]}利用SSR技术建立18个彩色棉品系的DNA指纹图谱。姜伟等^{[ 9]}利用ISSR标记, 对8个新疆品种(系) 构建指纹图谱。李武等^{[ 10]}利用SRAP标记, 对中国引入海岛棉以来培育的36个国内品种及20个国外品种进行了遗传多样性分析。潘兆娥等^{[ 11]}构建中棉所48的SSR数字指纹图谱。薛艳等^{[ 12]}建立新疆早熟棉品种SSR分子标记体系。李成奇等^{[ 13]}利用20对SSR核心引物构建了百棉系列棉花自交系品种(系)的DNA指纹图谱。孙宁等^{[ 14]}以2009年度参加黄河流域国家棉花区试的57份参试品种为材料进行指纹图谱分析。匡猛等^{[ 15]}采用SSR标记构建2008年中国3大棉区8个棉花主产省份32份棉花主栽品种的DNA指纹图谱并进行遗传多样性分析。赵亮等^{[ 16]}利用26对SSR引物构建12个品种DNA条形码编制。与其他分子标记相比, 以微卫星序列为基础的SSR标记在DNA指纹鉴定上显示了独特的优越性, SSR 标记数量丰富, 覆盖整个基因组, 揭示的多态性高, 以孟德尔方式遗传, 呈共显性^{[ 2]}。近年来, 基于SSR标记构建了水稻^{[ 17]}、小麦^{[ 18]}、玉米^{[ 19]}、烟草^{[ 20]}、大豆^{[ 21]}等主要农作物的DNA指纹图谱数据库。

本研究拟通过对51份新陆早常规棉花品种指纹图谱的建立及品种间遗传关系分析, 为种子管理经营单位提供棉花品种快速、准确、科学的鉴定方法, 保护育种者品种的知识产权和育种家的权益, 也为育种者提供亲本有效选配、品种合理种植的参考依据, 加快优质丰产新陆早棉花新品种的选育。

1 材料与方法

1.1 试验材料

2013年以前新疆审定的新陆早棉花常规品种51份, 统一由原品种选育单位提供(见附表)。SSR引物由华中农业大学作物遗传改良国家重点实验室提供, 引物合成、PCR试剂dNTPs、 Taq酶及电泳试剂等购自上海Sangon公司。

1.2 DNA提取及SSR检测

参照白静的棉花总DNA提取方法、SSR-PCR反应体系、PCR扩增程序、PAGE凝胶电泳及显色方法^{[ 22]}。

1.3 指纹的建立

根据PCR扩增产物在电泳凝胶上的相对位置, 对每对引物生成的不同基因型直接编号, 构建51份新陆早棉花品种的DNA分子指纹图谱。其中电泳结果采用0,1系统记录谱带位置, 某一扩增条带有带记为1, 无带记为0, 将每对引物在品种间扩增得到0,1 (二进制)数据转化成十进制数据, 该十进制数据代表每个引物扩增的结果, 则每40个引物组合的十进制数字串作为每一个品种的数字指纹。

1.4 统计分析

各引物对应不同基因型扩增, 计算位点多态性信息含量(polymorphism information content, PIC), 按公式PIC = 1-Σ P_i² 计算, 其中 P_i指第 i个等位变异出现的频率。采用UPGMA (unweighted pair-group method with arithmetic mean, 未加权平均数)法进行聚类分析, 并绘制树状聚类图。利用NTSYS-pcV 2.10软件中的Qualitative data计算品种间的遗传相似系数(Jaccard系数)。

2 结果与分析

2.1 SSR核心标记确定及标记信息分析

全面综合评价实验室已有的5000对引物, 选择多态性高、稳定性好、定位在棉花26条染色体上且分布均匀的标记引物91对, 在参试品种中进行筛选, 最终确定核心引物75对, 占检测引物的82.42%。

如表1所示, 75对SSR核心引物扩增检测到多态性基因型位点226个, 每个标记检测到的基因型位点数介于2~12个之间, 平均为3.01个。其中基因型数目大于平均数的标记47个, 引物编号分别为2、3、4、5、7、10、11、12、13、14、16、17、20、21、22、23、24、25、26、27、30、36、37、38、39、40、41、42、43、 46、48、50、52、54、56、57、60、61、64、65、66、68、69、70、71、73和74。图1所示, NAU3995标记在51份材料中扩增出等位基因数为4个。

表1所示, 标记间的多样性指数介于0.0799~ 0.8752之间, 平均多样性指数为0.6624, 其中多样性指数大于平均数的标记51个, 引物编号分别为1、2、3、4、5、7、8、9、10、11、12、13、14、15、16、20、21、22、23、24、25、26、27、30、31、32、34、36、37、38、39、41、42、43、44、45、46、48、50、53、54、56、57、60、61、62、64、65、66、70和71。表明这些标记具有较强鉴别品种的能力。

等位基因数和多样性指数均大于平均数的标记有40个, 如标记NAU3736, 等位基因数和基因型个数分别为5和12, 引物多态性信息值为0.8752。进一步证实挑选的核心标记引物有效性高, 标记的等位变异数多, 其多样性指数也高。

表1 75对SSR标记在51份新陆早棉花品种中的引物多态性信息 Table 1 Polymorphism information of 75 SSR loci in 51 Xinluzao cotton varieties

	Figure Option View Download New Window
	图1 标记NAU3995在51份新陆早棉花中电泳扩增等位基因特征Fig. 1 Alleles feature of marker NAU3995 in 51 Xinluzao cotton varieties

2.2 参试品种的指纹图谱分析

采用75对引物对51个新陆早棉花品种进行指纹分析, 其中21个新陆早棉花品种具有特征引物。在一定材料范围内, 特征引物是指某个品种具有明显区别与其他品种特有指纹信息的引物, 采用这对引物就可以直接区分该品种与其他品种。表2所示, 新陆早4号、新陆早10号、新陆早13、新陆早15、新陆早16、新陆早21、新陆早22、新陆早23、新陆早26、新陆早33、新陆早34、新陆早37、新陆早38、新陆早39、新陆早41、新陆早45和新陆早54分别具有1个特征引物, 新陆早3号、新陆早6号、新陆早40和新陆早47分别有3、3、6和2个特征引物。图2所示, BNL3257标记是新陆早41的特征引物。而NAU3736标记可以一批次分别鉴别出新陆早13、新陆早16、新陆早22、新陆早38、新陆早39和新陆早47。

表3所示, 其余30份材料需要引物组合来区别该品种与其他品种。例如, 在51份新陆早品种中区分新陆早1号, 第一步加标记HAU2022, 利用扩增出的特征带型(指纹信息)从51份材料中首先区分出来新陆早1号和新陆早3号, 第二步加新陆早3号特征引物NAU3995, 可明显区分开新陆早1号和新陆早3号, 即选用2对引物HAU2022和NAU3995组合就鉴别出新陆早1号。

依据表2和表3, 综合如表4所示, 最少选用40对特征引物和组合引物就可以完全区分开30份新陆早常规品种。因此, 可用这40对引物的指纹信息构建新陆早品种的指纹图谱。

	Figure Option View Download New Window
	图2 新陆早41的特征引物BNL3257电泳扩增图Fig. 2 Electrophoresis profiles of specific marker NAU1362 for Xinluzao 41

表2 新陆早棉花品种的特征引物 Table 2 Specific amplification primers for Xinluzao cotton varieties

表3 30份新陆早品种区分的组合引物 Table 3 Primer combinations for 30 Xinluzao cotton varieties

表4 利用40对引物构建51份新陆早棉花品种的指纹信息 Table 4 DNA fingerprinting of 51 Xinluzao cotton varieties using 40 primers

品种Variety	DNA指纹DNA fingerprinting
新陆早1号 Xinluzao 1	0-1-0-0-1-2-1-1-2-1-1-1-2-2-1-0-1-1-2-2-2-2-2-2-2-3-1-1-1-3-1-2-4-1-1-4-14-0-2-1
新陆早2号 Xinluzao 2	0-1-1-0-1-2-0-2-1-1-1-2-2-2-1-0-1-1-1-2-2-2-2-2-2-1-1-1-1-0-1-2-2-1-1-4-10-9-2-1
新陆早3号 Xinluzao 3	0-1-1-0-1-3-0-1-2-2-1-2-3-3-1-0-1-3-2-2-2-1-2-3-2-3-1-1-1-0-3-2-2-1-0-12-4-25-2-1
新陆早4号 Xinluzao 4	0-1-0-0-2-2-0-2-2-2-1-1-3-2-1-2-0-1-2-1-2-2-2-2-2-2-1-1-1-0-1-2-2-2-4-4-9-16-2-1
新陆早5号 Xinluzao 5	0-1-0-0-1-2-1-2-1-1-1-1-2-2-1-2-1-1-3-2-2-2-2-2-2-1-1-1-1-3-1-3-2-2-1-4-9-20-2-1
新陆早6号 Xinluzao 6	1-1-1-1-1-2-0-1-2-2-3-1-3-2-1-2-1-3-1-0-2-2-2-2-2-1-1-1-1-0-1-6-6-1-0-10-5-16-2-1
新陆早7号 Xinluzao 7	0-1-1-0-2-2-1-2-2-1-1-2-3-2-1-2-1-1-1-2-2-2-2-2-2-2-1-1-1-3-1-3-4-2-2-10-5-20-2-1
新陆早8号 Xinluzao 8	0-1-1-1-2-2-0-2-2-1-1-2-2-2-1-2-1-1-1-0-2-2-2-2-2-2-1-1-1-0-1-3-4-2-2-6-5-16-2-1
新陆早9号 Xinluzao 9	1-0-1-1-1-2-0-2-2-2-1-2-1-2-1-2-2-1-1-2-2-1-1-2-2-1-1-1-2-1-1-4-2-1-2-10-10-20-2-1
新陆早10号 Xinluzao 10	0-1-0-0-2-2-0-2-2-2-2-2-2-2-1-2-1-1-1-2-2-2-2-2-2-2-1-1-1-2-1-3-4-2-2-6-5-20-2-1
新陆早11 Xinluzao 11	0-1-1-1-1-2-1-1-2-1-1-2-2-2-1-0-2-1-1-1-2-3-2-0-1-2-2-1-0-0-1-4-2-0-0-9-0-20-3-1
新陆早12 Xinluzao 12	1-1-0-0-1-1-1-2-2-1-2-2-1-2-2-0-2-1-2-2-2-1-2-2-1-1-2-1-1-1-1-4-2-2-1-14-10-9-2-1
新陆早13 Xinluzao 13	1-1-0-0-1-2-0-2-2-1-1-2-1-2-1-0-2-1-1-2-2-3-2-2-2-1-1-1-2-1-2-4-2-2-2-10-6-11-1-2
新陆早15 Xinluzao 15	1-1-1-0-1-2-1-2-2-2-1-2-1-1-2-0-2-3-2-2-2-1-2-2-1-1-1-1-2-3-1-4-2-4-2-11-10-9-2-2
新陆早16 Xinluzao 16	0-1-0-0-1-2-1-2-2-2-1-1-2-2-1-2-1-1-1-1-2-2-1-2-2-1-2-1-1-3-1-4-2-1-2-9-5-26-2-0
新陆早17 Xinluzao 17	1-1-1-0-1-2-0-2-2-2-1-1-3-2-2-2-3-1-1-2-2-2-1-2-2-2-2-1-2-1-2-4-2-2-1-10-5-28-3-2
新陆早18 Xinluzao 18	1-1-0-1-1-2-0-2-2-2-1-2-3-2-2-1-2-3-1-2-2-1-2-2-2-2-2-1-2-3-2-4-2-1-2-6-6-0-3-2
新陆早19 Xinluzao 19	1-1-0-0-2-1-1-2-2-1-1-2-2-2-1-2-1-1-1-2-1-2-2-2-2-2-1-1-2-1-1-4-4-1-2-6-5-20-2-1
新陆早20 Xinluzao 20	1-1-0-1-1-2-1-2-2-2-1-1-2-2-1-2-1-1-1-1-2-2-2-2-2-1-2-1-2-3-1-4-2-1-0-14-5-28-3-1
新陆早21 Xinluzao 21	1-1-1-0-1-2-1-2-2-2-2-1-2-2-2-2-1-3-0-2-2-2-2-0-0-0-1-1-0-0-0-4-2-0-0-14-0-20-2-1
新陆早22 Xinluzao 22	1-1-0-0-1-2-1-2-2-1-2-1-2-2-1-1-2-3-1-2-2-2-1-2-2-1-1-1-1-3-2-4-2-1-1-6-6-18-1-2
新陆早23 Xinluzao 23	1-1-1-0-3-2-0-2-2-1-1-2-3-2-1-2-3-1-1-2-2-1-2-2-2-1-1-1-2-3-1-4-2-1-2-14-9-20-2-1
新陆早24 Xinluzao 24	1-1-0-0-1-1-1-2-2-2-1-2-2-2-1-2-2-1-2-1-2-1-1-2-1-1-1-1-1-3-1-4-2-2-2-6-10-20-2-1
新陆早25 Xinluzao 25	1-1-1-1-1-2-0-1-2-1-1-2-1-2-1-2-2-1-1-2-2-1-1-2-2-1-1-1-2-3-1-4-2-2-2-6-6-20-2-1
新陆早26 Xinluzao 26	1-1-0-1-2-2-1-2-2-2-1-1-1-2-2-2-1-2-2-2-2-2-2-1-2-1-1-1-1-3-1-4-2-0-1-6-9-20-2-1
新陆早27 Xinluzao 27	1-1-0-0-2-2-1-2-2-2-1-1-1-2-2-1-2-1-2-2-2-2-1-1-2-1-2-1-2-3-1-2-2-2-1-6-6-25-2-1
新陆早28 Xinluzao 28	0-1-1-0-1-1-1-2-2-2-1-2-2-2-1-2-2-1-2-1-2-1-1-2-1-1-1-1-1-3-1-4-2-2-2-6-10-28-2-1
新陆早29 Xinluzao 29	1-1-1-1-1-0-1-2-2-2-1-2-2-2-1-2-2-1-2-0-2-1-1-2-1-1-1-1-1-0-1-4-2-2-2-6-10-20-2-1
新陆早30 Xinluzao 30	1-1-1-1-1-1-1-2-2-2-1-2-2-2-1-2-2-1-2-1-2-1-1-2-1-1-1-1-1-3-1-4-2-2-2-6-10-20-2-1
新陆早31 Xinluzao 31	1-1-1-1-1-1-1-2-2-2-1-2-2-2-1-2-2-1-2-1-2-1-1-2-1-1-1-1-3-3-1-4-2-2-2-6-10-20-2-1
新陆早32 Xinluzao 32	1-1-0-1-2-3-0-2-2-2-1-2-2-2-1-1-0-1-1-0-2-1-1-2-2-1-2-1-1-0-2-4-2-1-0-6-5-16-2-0
新陆早33 Xinluzao 33	1-1-0-0-1-3-1-2-2-1-2-1-2-2-1-1-0-3-1-2-2-2-1-2-2-1-1-1-3-0-2-4-1-2-1-6-6-16-1-1
新陆早34 Xinluzao 34	1-1-0-0-2-1-1-2-2-1-1-2-2-2-1-2-1-1-1-2-1-2-2-2-2-2-1-1-2-1-1-4-4-1-2-6-12-20-2-1
新陆早35 Xinluzao 35	1-1-0-0-1-3-0-2-2-2-1-1-2-2-1-1-2-3-1-2-2-2-2-2-2-1-1-1-3-3-1-4-2-1-2-5-14-2-1-1
新陆早36 Xinluzao 36	1-1-0-0-2-3-1-2-2-1-1-2-2-2-1-2-0-1-1-2-0-1-0-0-2-0-1-1-1-0-1-0-4-0-0-5-0-16-2-1
新陆早37 Xinluzao 37	1-1-1-1-1-3-1-2-2-2-1-2-1-2-1-0-1-3-1-2-2-1-1-2-2-2-1-2-3-1-1-3-2-1-2-6-9-9-2-2
新陆早38 Xinluzao 38	1-0-1-0-1-1-0-2-2-2-2-2-3-2-2-1-2-3-1-2-2-1-2-2-2-1-1-1-2-1-1-4-2-1-2-10-10-10-2-1
新陆早39 Xinluzao 39	1-1-0-0-1-1-1-2-2-2-1-2-3-2-1-2-0-3-0-2-2-1-1-2-1-1-1-1-1-0-1-4-2-0-0-6-10-24-2-1
新陆早40 Xinluzao 40	1-1-1-1-1-2-0-3-2-1-1-3-3-2-3-2-1-3-3-2-2-1-3-2-2-1-1-1-3-3-1-4-2-3-0-6-7-20-2-1
新陆早41 Xinluzao 41	1-1-0-1-1-2-0-2-2-3-2-2-3-2-2-2-0-3-1-2-2-1-0-2-0-0-1-1-2-0-2-4-2-0-0-6-10-16-2-1
新陆早42 Xinluzao 42	1-1-0-0-2-2-1-2-2-1-1-2-2-2-1-1-1-3-2-2-2-2-1-2-2-1-1-1-1-3-2-4-2-2-0-6-5-0-2-2
新陆早45 Xinluzao 45	1-1-0-0-1-2-0-2-2-1-1-2-2-2-1-3-2-3-1-2-2-1-1-2-2-1-1-1-1-1-1-4-2-1-1-6-6-2-3-2
新陆早46 Xinluzao 46	1-1-0-1-2-2-1-2-2-2-2-1-3-2-1-1-1-3-2-2-2-2-2-2-2-2-1-1-1-1-1-3-2-2-3-10-5-2-2-1
新陆早47 Xinluzao 47	1-1-1-1-1-1-1-2-2-1-1-2-3-2-2-2-2-3-1-1-2-1-2-2-1-1-3-1-1-1-1-4-2-1-3-11-6-29-2-1
新陆早48 Xinluzao 48	1-1-0-0-2-2-1-2-2-1-2-1-2-2-1-2-1-3-1-2-2-2-1-2-2-1-1-1-1-3-1-4-2-2-2-6-5-28-2-1
新陆早49 Xinluzao 49	1-1-0-0-1-2-0-2-2-2-1-2-1-1-2-2-2-3-1-2-2-1-2-2-2-1-1-1-1-3-2-4-2-1-2-10-14-20-3-1
新陆早50 Xinluzao 50	1-1-0-0-1-2-1-2-2-2-2-2-1-2-2-2-2-3-1-2-2-1-2-2-1-2-1-1-2-1-2-4-2-1-2-6-10-20-2-1
新陆早51 Xinluzao 51	1-1-0-0-2-2-1-2-2-1-1-2-2-2-1-1-1-3-2-2-2-2-1-2-2-1-1-1-1-3-2-4-2-2-2-6-5-2-2-2
新陆早52 Xinluzao 52	1-1-1-1-1-1-1-1-3-2-1-1-3-1-1-2-3-2-2-1-2-3-1-2-0-1-1-1-1-3-1-6-2-2-2-15-11-24-1-1
新陆早53 Xinluzao 53	1-1-0-0-1-2-0-2-2-1-2-1-2-2-1-1-1-1-1-2-2-2-1-2-2-1-1-1-3-1-2-4-1-2-1-15-6-16-1-2
新陆早54 Xinluzao 54	0-1-0-0-1-2-1-2-2-3-1-2-2-3-1-2-2-1-2-2-2-3-2-2-1-2-3-1-1-1-2-1-4-2-2-1-15-10-20-2-3

40 primers: NAU1071-BNL598-BNL1421-NAU5099-NAU1190-NAU934-CIR43-NAU1362-BNL3255-BNL3257-BNL3442-MUSS281- NAU3778-TMB71-NAU5499-CIR307-NAU3095-NAU3110-BNL3948-BNL119-BNL3646-BNL1655-NAU3732-NAU2631-BNL1521-HAU2022- MGHES44-CIR170-DPL238-NAU2437-MUSS95-BNL663-BNL2486-Gh277-BNL3173-NAU3995-BNL1317-NAU3736-NAU1042-MUCS101.

表4 利用40对引物构建51份新陆早棉花品种的指纹信息 Table 4 DNA fingerprinting of 51 Xinluzao cotton varieties using 40 primers

2.3 遗传多样性分析

根据SSR分子标记统计不同基因型数据结果, 利用NTSYS-pcV2.10软件, 采用Jaccard系数计算51个新陆早常规品种遗传相似系数, 得到相似系数矩阵。结果表明, 51个新陆早品种遗传相似系数变化范围是0.4269~0.9873, 平均值为0.7071, 遗传相似系数较高, 表明新陆早棉花品种之间遗传多样性较狭窄。遗传相似系数最大的品种是新陆早30和新陆早31, 表明二者的遗传差异较小; 遗传相似系数最小是新陆早15和新陆早32, 表明两者遗传差异较大, 亲缘关系较远。

根据品种遗传相似系数矩阵, 对品种聚类( 图3), 在相似系数为0.4619处可以将51个新陆早棉花品种分为4大类, A类包括新陆早13个棉花品种, B类包括29个, C类包括2个, D类包括7个, 其中B类在遗传相似系数0.4778处又可分为2个亚类, 即B1与B2, B1包含16个品种, B2包含13个品种。对比各新陆早品种系谱可明显看出, A类以石河子棉花研究所(有新陆早2号、新陆早5号、新陆早7号、新陆早8号、新陆早10号、新陆早19、新陆早36、新陆早46)及农七师农业科学研究所(有新陆早3号、新陆早4号、新陆早6号)选育的品种为主, 其亲本多含有前苏联早熟陆地面血缘, 其中新陆早1号、新陆早6号、新陆早7号、新陆早8号、新陆早36在北疆早熟棉区作为主载品种推广面积较大。

B类以农七师农业科学研究所(新陆早9号、新陆早13、新陆早15、新陆早16、新陆早23、新陆早25、新陆早31、新陆早35、新陆早37、新陆早38、新陆早39、新陆早47和新陆早49)及农五师地区(新陆早11、新陆早12、新陆早37、新陆早54)所选育品种为主, 新陆早系列品种中, 优质中长绒或抗病性较好的品种主要集中在B类中。其亲本多含有贝尔斯诺、爱子棉、中棉所12、中棉所17及辽棉系列的资源, 且通过审定的中长绒品种品种主要都聚类在B2亚类中(有新陆早24、新陆早25、新陆早28、新陆早29、新陆早30、新陆早31、新陆早35、新陆早38、新陆早39、新陆早40、新陆早47、新陆早49)。其中新陆早12和新陆早13在北疆早熟棉区作为主载品种推广。

C类中的包含新陆早26、新陆早27, 其中新陆早26在北疆早熟棉区推广面积较大。

D类新陆早品种主要由新疆农垦科学院棉花研究所(有新陆早22、新陆早32、新陆早33、新陆早42、新陆早48、新陆早51、新陆早53)选育, 其中新陆早33和新陆早48在北疆早熟棉区为主载品种。新疆农垦科学院棉花研究所选育的新陆早45的亲本之一来源于新陆早13, 在分子标记多样性聚类分析中, 与新陆早13聚为一类和系谱描述高度吻合。

	Figure Option View Download New Window
	图3 51个新陆早棉花品种的聚类分析图Fig. 3 Clustering analysis of 51 Xinluzao cotton varieties

3 讨论

随着棉花全基因组测序迅速进展, 构建高通量、准确性高, 易操作的全基因组覆盖DNA指纹库将越来越受到重视。一种理想的分子标记应具有以下特点, 即标记引物的多态性高、重复性和稳定性好; 带型清晰, 易统计; 染色体上均匀分布; 共显性; 开发和使用成本低。DNA指纹技术的发展经历3个阶段, 第1代是以Southern杂交为基础的RFLP, 第2代是以PCR为基础的DNA指纹标记(如RAPD、AFLP、SSR、SCAR、ISSR等), 第3代是以单核苷酸多态性为基础的SNP^{[ 23]}。近些年, 分子标记开始应用于各物种的鉴定, 国际植物品种权保护组织(UPOV)在BMT测试指南草案中已将构建DNA指纹数据库的标记确定为SSR和SNP^{[ 24]}。随着第2代测序技术的发展, 使品种基因组DNA测序和SNP全面分析成为可能, 从而大大提升了品种识别和鉴定精度, 但检测成本较高, 有时会因检测尺度的过度精细, 把品种内的差异误判成品种间的差异^{[ 25]}。因此, SNP高额的投入使其未能得到广泛应用^{[ 26]}, 其中SSR标记技术比较成熟, 成为当前各个作物构建指纹数据库的首选技术。本研究采用SSR技术构建了新陆早常规品种指纹图谱, 并分析和研究了品种的遗传多样性和亲缘关系。

本试验的聚丙烯凝胶电泳采用变性胶, 而薛艳等^{[ 12]}研究42份新疆陆地早熟棉指纹图谱构建与品种鉴别时采用9%非变性聚丙烯酰胺凝胶电泳分离方法, 曲鲁江等^{[ 27]}报道的非变性凝胶中条带过于复杂, 出现的非特异性条带极大地影响了分析带形的准确性, 使试验结果的误差加大, 变性胶在利用银染检测微卫星产物时条带清晰, 带型分析准确性更高, 结果更加可靠。

本研究以75对核心引物SSR标记扩增检测到多态性基因型位点226个, 每个标记检测到的基因型位点数在2~12之间, 平均为3.01个, 其中等位基因型数目大于平均数的标记47个; 标记间的多样性指数介于0.0799~0.8752之间, 平均数为0.6624, 多样性指数大于平均数的标记51个。张玉翠等^{[ 2]}用32个主栽品种, 40对引物, 共扩增出161种多态性基因型, 引物多态信息量(PIC)为0.2989~0.7585, 平均为0.5407; 艾先涛等^{[ 28]}用94份新疆陆地棉品种, 54对具有稳定多态性的引物, 共检测出153个多态性位点, 每对引物的等位变异为2~6个, 平均为2.93个; 引物多态信息含量(PIC)为0.0430~0.6640, 平均为0.3831。与其相比, 本研究筛选的引物标记是高效的。

51个新陆早棉花品种采用75对引物进行指纹分析, 其中21个品种具有特征引物, 获得特征引物为23个, 其中NAU3736标记可以一批次分别鉴别出6个品种, 而引物NAU3736在表1中, 等位基因数、基因型个数和PIC值分别为5、12、0.8752, 结果表明引物多态性越丰富且特征谱带数量越多的标记引物其鉴别品种能力越强, 可以首选作为构建新陆早棉花品种指纹图谱的标记。其余30份材料没有获得特征引物, 采用引物组合来区别该品种与其他品种, 共选用组合引物34个。最终统计, 至少确定40对特征引物和组合引物就可以完全区分开新陆早常规品种。以上研究表明, 特征引物在一定材料范围内鉴定是最快捷、易见的, 但是只有部分品种具有特征引物, 随着鉴定材料数量的扩大, 需要采用组合引物鉴定, 可大幅度提高鉴别能力。本研究中全面、系统构建的新陆早51个品种指纹图谱, 每一个品种的指纹都是唯一的, 具有很强的特异性, 此数据库可以用于新陆早棉花品种初级数据库。与赵亮^{[ 16]}等的研究具有明显差异, 本试验选用新疆2013年前审定的早熟陆地棉品种全套完整材料, 75对核心标记均匀分布于棉花26条染色体且较高多态性, 结合特征引物和组合引物方法构建品种指纹。

51个新陆早品种遗传相似系数矩阵和品种聚类表明, 新陆早棉花品种之间遗传多样性较狭窄, 这与艾先涛等^{[ 28]}的研究结果相似, 说明新疆陆地棉品种间遗传关系相对简单, 品种的遗传基础相对狭窄, 品种遗传组分差异较小, 总体上遗传多样性不够丰富。在相似系数0.4619处将51个新陆早品种分为4大类, 其中每类包括的新陆早品种基本是来源于一个地区育种单位, 例如, A类包括石河子棉花研究所和农七师农业科学研究所选育品种, B类包括农七师农业科学研究所和农五师农业科学研究所选育品种, C类是2个公司选育品种, D类包括新疆农垦科学院棉花研究所选育品种, 具有相同来源的品种最先聚为一起; 遗传相似系数最大的品种是新陆早30和新陆早31, 表明其遗传差异较小, 结合表3显示, 新陆早30和新陆早31分别用标记BNL119、BNL1421组合和标记DPL238、BNL2486、NAU3995、CIR170、BNL1317组合引物鉴别, 因此遗传差异较小的品种使用组合引物标记鉴别才更为准确。以上研究结果与Liu等^{[ 29]}的观点一致, 即SSR聚类结果与地理分布一致, 遗传相似系数揭示种群之间的正相关关系; 从附表看出, 新陆早9号、新陆早16、新陆早25、新陆早31和新陆早39的亲本均来源于贝尔斯诺系统, 因此B类品种主要亲本来源有贝尔斯诺、爱字棉系统、中棉所12、中棉所17、辽棉系列, 此系列选育品种数量最多、棉花纤维品质较优, 说明聚类结果和品种根据实际系谱来源分类是一致的, 这与张玉翠等^{[ 2]}的研究结果一致, 即遗传相似系数聚类分析结果基本上反映了品种之间的亲缘关系。新疆北疆早熟陆地棉品种间分子聚类结果与品种本身遗传系谱背景高度吻合, 符合品种本身的真实特性。具有相近表型性状、相同的遗传背景和相同育种单位选育的品种聚在同一类群中。说明北疆不同育种单位在早熟棉选育过程中积累形成了各自的特点及优势, 表现在各育种单位在基础种质资源上确立了其特定的选育方向。

今后应将形态信息与分子数据结合一起形成指纹图谱, 使品种保护信息更加全面、准确。目前已在油菜^{[ 30]}、大豆^{[ 31]}构建分子身份证, 在甜瓜^{[ 32]}上建立指纹图谱QR编码, 在水稻^{[ 33]}上提出基于SSR分子指纹和商品信息构建品种身份证的新思路。因此, 棉花DNA指纹鉴定技术发展将不断完善, 将棉花的指纹信息编码结合品种的基本商品信息和特异基因信息构建棉花品种身份证。

4 结论

利用SSR分子标记技术, 通过40对特征引物和组合引物构建了新陆早51份棉花品种指纹图谱, 将51个新陆早品种聚为4大类型, 遗传多样性较狭窄。构建的指纹图谱可以作为新陆早棉花品种初级分子标记指纹数据库。

致谢: 本研究的试验材料来自各品种育种单位提供的原原种, 主要由新疆农垦科学院李保成和余渝, 石河子棉花研究所孔宪良、黄顶元及秦江鸿, 农七师农科所李家胜和赵富强, 农五师农科所曹阳提供; 分子标记实验由作物遗传改良国家重点实验室林忠旭教授精心设计指导, 同时得到黄聪、郭欢乐、王夏青等研究生的帮助, 在此表示真诚的感谢。

附表新陆早棉花品种的信息 Supplementary table Informations of Xinluzao cotton varieties

品种 Variety	系谱 Pedigree	品种选育单位 Released institution
新陆早1号 Xinluzao 1	农垦5号选系722 Line 722 from Nongken 5	下野地实验站 Xiayedi Experimental Station
新陆早2号 Xinluzao 2	6902×中棉所4号 6902×Zhongmiansuo 4	农八师农业科学研究所 Nongbashi Agricultural Institute
新陆早3号 Xinluzao 3	(79-73W×爱子棉)×荆州4588 (79-73W×Aizimian) ×Jinzhou 4558	农七师农业科学研究所 Nongqishi Agricultural Institute
新陆早4号 Xinluzao 4	(66-241×灃74-47)F₁×岱70 (66-241× Li 74-47)F₁×Dai 70	农七师农业科学研究所 Nongqishi Agricultural Institute
新陆早5号 Xinluzao 5	(347-2×科遗181)F₁×(83-2-3+陕1155) (347-2×Keyi 181)F₁×(83-2-3+Shaan 1155)	农八师农业科学研究所 Nongbashi Agricultural Institute
新陆早6号 Xinluzao 6	85-174×贝尔斯诺 85-174×Beiersinuo	农七师农业科学研究所 Nongqishi Agricultural Institute
新陆早7号 Xinluzao 7	3347×塔什干2号 3347×Tashigan 2	农八师农业科学研究所 Nongbashi Agricultural Institute
新陆早8号 Xinluzao 8	(抗V.Wx×早1号)F₁辐射 (Kang V.Wx×Zao 1) F₁ radiation	农八师农业科学研究所 Nongbashi Agricultural Institute
新陆早9号 Xinluzao 9	(新陆早6号×贝尔斯诺)×中棉所17 (Xinluzao 6×Beiersinuo)×Zhongmiansuo17	农七师农业科学研究所 Nongqishi Agricultural Institute
新陆早10号 Xinluzao 10	(黑山棉×02II)F₂×中棉所12 (Heishanmian×02II)F₂×Zhongmiansuo 12	农八师农业科学研究所 Nongbashi Agricultural Institute
新陆早11 Xinluzao 11	豫早202系选 Yuzao 202 breeding line	博乐种子站 Bole Seed Banks
新陆早12 Xinluzao 12	辽95-25病圃系选 Liao 95-25 breeding line from disease nursery	农五师农业科学研究所 Nongwushi Agricultural Institute
新陆早13 Xinluzao 13	83-14 ×(中无5601+1639) 83-14 ×(Zhongwu 5601+1639)	农七师农业科学研究所 Nongqishi Agricultural Institute
新陆早15 Xinluzao 15	JW低酚×中棉所12 JW low gossypol × Zhongmiansuo 12	农七师农业科学研究所 Nongqishi Agricultural Institute
新陆早16 Xinluzao 16	早熟鸡脚×贝尔斯诺 Zaoshujijiao×Beiersinuo	农七师农业科学研究所 Nongqishi Agricultural Institute
新陆早17 Xinluzao 17	9908系选 9908 breeding line	新疆农业科学院 Xinjiang Academy of Agricutural Sciences
新陆早18 Xinluzao 18	69118系选 69118 breeding line	新疆农业科学院 Xinjiang Academy of Agricutural Sciences
新陆早19 Xinluzao 19	91-2×900	农八师农业科学研究所 Nongbashi Agricultural Institute
新陆早20 Xinluzao 20	新陆早16 (97-185)病圃系选 Xinluzao 16 (97-185) breeding line from disease nursery	150团 150 Group
新陆早21 Xinluzao 21	新陆早8号抗病变异株 Xinluzao 8 variation plant with resistant disease	富依德公司 Fuyide Company
新陆早22 Xinluzao 22	优系451×新陆早6号 Elite line 451×Xinluzao 6	新疆农垦科学院 Xinjiang Land Reclamation Academy of Sciences
新陆早23 Xinluzao 23	中棉所27变异 Zhongmiansuo 27 variation plant	万氏种业 Wanshi Seed Industry
新陆早24 Xinluzao 24	中长绒品系7047×C6524 Middle length fiber line 7047×C6524	康地种业 Kangdi Seed Industry
新陆早25 Xinluzao 25	[(系5×贝尔斯诺)×晋14] ×中棉所17 [(Xi 5×Beiersinuo)×Jin 14] ×Zhongmiansuo 17	农七师农业科学研究所 Nongqishi Agricultural Institute
新陆早26 Xinluzao 26	新陆早8号(1304)变异株 Xinluzao 8 (1304) variation plant	天合种业 Tianhe Seed Industry
新陆早27 Xinluzao 27	早熟抗病7147×贝尔斯诺 Zaoshukangbing 7147×Beiersinuo	康地种业 Kangdi Seed Industry
新陆早28 Xinluzao 28	85-57×(贝尔斯诺+西南农大抗病, 优质, 丰产材料混合花粉) 85-57×(Beiersinuo +Southwest Agricultural University mixture pollen with resistant disease, high quality and high yield)	惠远公司 Huiyuan Company
新陆早29 Xinluzao 29	中棉所、辽棉所、河南省农科院等单位引试验品种(系)混播选育 Testing cotton varieties (or lines) mixed seeding breeding from Institute of Cotton Research of CAAS, Institute of Economic Crop of LAAS, and Henan Academy of Agricultural Sciences	金博种业 Jinbo Seed Industry
新陆早30 Xinluzao 30	中棉所、辽棉所、河南省农科院等单位引试验品种(系)混播选育 Testing cotton varieties (or lines) mixed seeding breeding from Institute of Cotton Research of CAAS, Institute of Economic Crop of LAAS, and Henan Academy of Agricultural Sciences	金博种业 Jinbo Seed Industry
新陆早31 Xinluzao 31	(新陆早6号×贝尔斯诺) F₁×岱子棉 (Xinluzao 6×Beiersinuo) F₁×Daizimian	万氏种业 Wanshi Seed Industry
新陆早32 Xinluzao 32	拉玛干77变异株病圃系选 Lamagan 77 variant plant from disease nursery	新疆农垦科学院 Xinjiang Land Reclamation Academy of Sciences
新陆早33 Xinluzao 33	石选87变异株病圃系选 Shixuan 87 variant plant from disease nursery	新疆农垦科学院 Xinjiang Land Reclamation Academy of Sciences
新陆早34 Xinluzao 34	早熟系97-65×7003 Zaoshuxi 97-65×7003	康地种业 Kangdi Seed Industry
新陆早35 Xinluzao 35	[(新陆早3号×中2621) ×抗35]×97-185 [(Xinluzao 3×Zhong 2621) ×Kang 35]×97-185	农七师农业科学研究所 Nongqishi Agricultural Institute
新陆早36 Xinluzao 36	新陆早8号×抗病品系BD103 Xinluzao 8×Resistant disease line BD103	农八师农业科学研究所 Nongbashi Agricultural Institute
新陆早37 Xinluzao 37	(辽83421×系5)×(辽9001+系5+自育90-5) (Liao 83421×Line 5)×(Liao 9001+ Line 5+ self-breeding 90-5)	农五师农业科学研究所 Nongwushi Agricultural Institute
新陆早38 Xinluzao 38	[(92-226×中6331)×中17]×97-145 [(92-226×Zhong 6331)×Zhong 17]×97-145	农七师农业科学研究所 Nongqishi Agricultural Institute
新陆早39 Xinluzao 39	(新陆早4号×贝尔斯诺) ×岱字棉 (Xinluzao 4×Beiersinuo)×Daizimian	万氏种业 Wanshi Seed Industry
新陆早40 Xinluzao 40	97-185×(D256×sw2) F₂	新疆农垦科学院 Xinjiang Land Reclamation Academy of Sciences
新陆早41 Xinluzao 41	高代材料17-79系选 17-79 line from high generation	富全新科 Fuquanxinke
新陆早42 Xinluzao 42	新陆早10×97-6-9 Xinluzao 10×97-6-9	新疆农垦科学院 Xinjiang Land Reclamation Academy of Sciences
新陆早45 Xinluzao 45	新陆早13×9941 Xinluzao 13×9941	新疆农垦科学院 Xinjiang Land Reclamation Academy of Sciences
新陆早46 Xinluzao 46	系9×抗病822 Line 9×Resistant disease 822	农八师农业科学研究所 Nongbashi Agricultural Institute
新陆早47 Xinluzao 47	(中17×9001)×97-185 (Zhong 17×9001)×97-185	农七师农业科学研究所 Nongqishi Agricultural Institute
新陆早48 Xinluzao 48	石选87×优系604 Shixuan 87× Elite line 604	惠远公司 Huiyuan Company
新陆早49 Xinluzao 49	9765 ×新陆早16 9765 ×Xinluzao 16	农七师农业科学研究所 Nongqishi Agricultural Institute
新陆早50 Xinluzao 50	新陆早13×Y-605 Xinluzao 13 ×Y-605	新疆农业科学院 Xinjiang Academy of Agricutural Sciences
新陆早51 Xinluzao 51	新陆早10×97-6-9×垦0074 Xinluzao 10×97-6-9×Ken 0074	新疆农垦科学院 Xinjiang Land Reclamation Academy of Sciences
新陆早52 Xinluzao 52	硕丰1号优系4004×早28选系 Shuofeng 1 4004×Zao 28 breeding line	硕丰种业 Shuofeng Seed Industry
新陆早53 Xinluzao 53	石选87×新陆早9号 Shixuan 87×Xinluzao 9	新疆农垦科学院 Xinjiang Land Reclamation Academy of Sciences
新陆早54 Xinluzao 54	新陆早11×中棉所12 Xinluzao 11×Zhongmiansuo 12	金宏祥高科农业股份有限公司 Jinhongxiang High-Tech Agriculture Co.

附表新陆早棉花品种的信息 Supplementary table Informations of Xinluzao cotton varieties

The authors have declared that no competing interests exist.

作者已声明无竞争性利益关系。

参考文献

View Option

[1]	张玉翠. 我国棉花品种SSR指纹数据库的初步构建. 中国农业科学院研究生院硕士论文, 北京, 2012. pp1-8 Zhang Y C. Construction a Primary SSR Fingerprint Database for Cotton Varieties. MS Thesis of Graduate School of Chinese Academy of Agricultural Sciences, Beijing, China, 2012. pp1-8 (in Chinese with English abstract) [本文引用:1]
[2]	张玉翠, 杨伟华, 匡猛, 许红霞, 王延琴, 周大云, 冯新爱, 苏畅. 32个棉花主栽品种DNA指纹图谱构建及遗传多样性分析. 棉花学报, 2012, 24: 120-126 Zhang Y C, Yang W H, Kuang M, Xu H X, Wang Y Q, Zhou D Y, Feng X A, Su C. Construction of DNA fingerprinting and analysis of genetic diversity with SSR markers for 32 leading cotton varieties. Cotton Sci, 2012, 24: 120-126 (in Chinese with English abstract) [本文引用:4] [CJCR: 1.462]
[3]	Zhang H Y, Wang H, Guo S, Yi R, Guo Y G, Yi Q W, Yong X. Identification and validation of a core set of microsatellite markers for genetic diversity analysis in watermelon, Citrullus lanatus Thunb. Matsum. & Nakai. Euphytica, 2012, 186: 324-342 [本文引用:1] [JCR: 1.643]
[4]	郭旺珍, 张天真, 潘家驹, 何金龙. 我国棉花主栽品种的RAPD指纹图谱研究. 农业生物技术学报, 1996, 4: 129-134 Guo W Z, Zhang T Z, Pan J J, He J L. Analysis of RAPD fingerprinting on main cotton varieties in China. J Agric Biotechnol, 1996, 4: 129-134 (in Chinese with English abstract) [本文引用:1]
[5]	Multani D S, Lyon B R. Genetic fingerprinting of Australian cotton varietys with RAPD markers. Genome, 1995, 38: 1005-1008 [本文引用:1] [JCR: 1.668]
[6]	Iqbal M J, Aziz N, Saeed N A, Zafar Y, Malik K A. Genetic diversity evaluation of some elite cotton varieties by RAPD analysis. Theor Appl Genet, 1997, 94: 139-144 [本文引用:1] [JCR: 3.658]
[7]	武耀廷, 张天真, 郭旺珍, 殷剑美. 陆地棉品种SSR标记的多态性及用于杂交种纯度检测的研究. 棉花学报, 2001, 13: 131-133 Wu Y T, Zhang T Z, Guo W Z, Yin J M. Detecting polymorphism among upland cotton (Gossypium hirsutum L. ) cultivars and their roles in seed purity of hybrids with SSR markers. Cotton Sci, 2001, 13: 131-133 (in Chinese with English abstract) [本文引用:1] [CJCR: 1.462]
[8]	马轩, 杜雄明, 孙君灵. 18个彩色棉品系的SSR指纹分析. 植物遗传资源学报, 2003, 4: 305-310 Ma X, Du X M, Sun J L. SSR fingerprinting analysis on 18 colored cotton lines. J Plant Genet Resourc, 2003, 4: 305-310 (in Chinese with English abstract) [本文引用:1] [CJCR: 1.1628]
[9]	姜伟, 李雪源, 何觉民, 陆建农. 新疆8个棉花品种(系)的指纹图谱分析. 华北农学报, 2007, 22: 89-92 Jiang W, Li X Y, He J M, Lu J N. Fingerprinting analysis for 8 cultivars (lines) of Xinjiang. Acta Agric Boreali-Sin, 2007, 22: 89-92 (in Chinese with English abstract) [本文引用:1] [CJCR: 0.951]
[10]	李武, 倪薇, 林忠旭, 张献龙. 海岛棉遗传多样性的SRAP标记分析. 作物学报, 2008, 34: 893−898 Li W, Ni W, Lin Z X, Zhang X L. Genetic diversity analysis of sea-island cotton cultivars using SRAP markers. Acta Agron Sin, 2008, 34: 893−898 (in Chinese with English abstract) [本文引用:1] [CJCR: 1.667]
[11]	潘兆娥, 王希文, 孙君灵, 周忠丽, 贾银华, 何守朴, 王杰, 王立如, 庞保印, 杜雄明. 中棉所48的SSR数字指纹图谱的构建. 中国农学通报, 2010, 26(7): 31-35 Pan Z E, Wang X W, Sun J L, Zhou Z L, Jia Y H, He S P, Wang J, Wang L R, Pang B Y, Du X M. Construction of digital fingerprint of Zhongmiansuo 48. Chin Agric Sci Bull, 2010, 26(7): 31-35 (in Chinese with English abstract) [本文引用:1]
[12]	薛艳, 李保成, 张新宇, 沙红, 孙杰. 新疆早熟棉品种SSR分子标记体系的建立. 石河子大学学报(自然科学版), 2011, 29(1): 1-5 Xue Y, Li B C, Zhang X Y, Sha H, Sun J. Optimization of amplification reaction system and construction of DNA fingerprinting of local upland cotton variety in Xinjiang. J Shihezi Univ (Nat Sci), 2011, 29(1): 1-5 (in Chinese with English abstract) [本文引用:2] [CJCR: 0.4744]
[13]	李成奇, 王晓芸, 张晓飞, 王清连. 百棉系列棉花自交系品种(系)SSR指纹图谱构建. 棉花学报, 2011, 23: 228-234 Li Ch Q, Wang X Y, Zhang X F, Wang Q L. SSR fingerprinting establishment of baimian series cotton inbred varieties (lines). Cotton Sci, 2011, 23: 228-234 [本文引用:1] [CJCR: 1.462]
[14]	孙宁, 杨付新, 付小琼, 王秀玲, 刘逢举, 李亚丽, 汤磊鹏. 国家棉花区试新品种的SSR指纹图谱分析. 棉花学报, 2011, 23: 279-283 Sun N, Yang F X, Fu X Q, Wang X L, Liu F J, Li Y L, Tang L P. SSR marker fingerprinting of cotton varieties in national regional trails. Cotton Sci, 2011, 23: 279-283 (in Chinese with English abstract) [本文引用:1] [CJCR: 1.462]
[15]	匡猛, 杨伟华, 许红霞, 王延琴, 周大云, 冯新爱. 中国棉花主栽品种DNA指纹图谱构建及SSR标记遗传多样性分析. 中国农业科学, 2011, 44: 20-27 Kuang M, Yang W H, Xu H X, Wang Y Q, Zhou D Y, Feng X A. Construction of DNA fingerprinting and analysis of genetic diversity with SSR markers for cotton major cultivars in China. Sci Agric Sin, 2011, 44: 20-27 (in Chinese with English abstract) [本文引用:1] [CJCR: 1.889]
[16]	赵亮, 蔡彩平, 梅鸿献, 郭旺珍. 用于区别不同棉花品种基因组特征的微卫星位点筛选. 作物学报, 2012, 38: 1810-1817 Zhao L, Cai C P, Mei H X, Guo W Z. Screening of microsatellite loci for identifying genome barcoding of cotton cultivars. Acta Agron Sin, 2011, 44: 20-27 (in Chinese with English abstract) [本文引用:2] [CJCR: 1.667]
[17]	程保山, 万志兵, 洪德林. 35个粳稻品种SSR指纹图谱的构建及遗传相似性分析. 南京农业大学学报, 2007, 30(3): 1-8 Cheng B S, Wan Z B, Hong D L. Establishment of SSR fingerprint map and analysis of genetic similarity among 35 varieties in japonica rice. J Nanjing Agric Univ, 2007, 30(3): 1-8 (in Chinese with English abstract) [本文引用:1] [CJCR: 0.916]
[18]	李根英, Dreisigacker S, Warburton M L, 夏先春, 何中虎, 孙其信. 小麦指纹图谱数据库的建立及SSR分子标记试剂盒的研发. 作物学报, 2006, 32: 1771-1778 Li G Y, Dreisigacker S, Warburton M L, Xia X C, He Z H, Sun Q X. Development of a fingerprinting database and assembling an SSR reference kit for genetic diversity analysis of wheat. Acta Agron Sin, 2006, 32: 1771-1778 (in Chinese with English abstract) [本文引用:1] [CJCR: 1.667]
[19]	赵久然, 王凤格, 郭景伦, 陈刚, 廖琴, 孙世贤, 陈如明, 刘龙洲. 中国玉米新品种DNA指纹库建立系列研究. 玉米科学, 2003, 11(2): 3-5 Zhao J R, Wang F G, Guo J L, Chen G, Liao Q, Sun S X, Chen R M, Liu L Z. Series of research on establishing DNA fingerprinting pool of Chinese new maize varieties. J Maize Sci, 2003, 11(2): 3-5 (in Chinese with English abstract) [本文引用:1] [CJCR: 0.965]
[20]	徐军, 刘艳华, 任民, 牟建民, 张兴伟, 陈雅琼, 王志德. 普通烟草种质资源的SSR标记与指纹图谱分析. 中国烟草科学, 2011, 32(2): 62-65 Xu J, Liu Y H, Ren M, Mou J M, Zhang X W, Chen Y Q, Wang Z D. SSR fingerprint map analysis of tobacco germplasms. Chin Tob Sci, 2011, 32(2): 62-65 (in Chinese with English abstract) [本文引用:1] [CJCR: 1.2747]
[21]	田清震, 盖钧镒, 喻德跃, 吕慧能, 贾继增. 我国野生大豆与栽培大豆AFLP指纹图谱研究. 中国农业科学, 2001, 34: 465-468 Tian Q Z, Gai J Y, Yu D Y, Lü H N, Jia J Z. AFLP fingerprint analysis of G. soja and G. max in China. Sci Agric Sin, 2001, 34: 465-468 (in Chinese with English abstract) [本文引用:1] [CJCR: 1.889]
[22]	白静. 杂交棉品种SSR核心引物的筛选与真实性和纯度鉴定. 华中农业大学硕士论文, 湖北武汉, 2011, pp45-50 Bai J. Screening of SSR Core Primer and the Identification of Genuineness and Purity on Hybrid Cotton. MS Thesis of Huazhong Agricultural University, Wuhan, China, 2011. pp45-50 (in Chinese with English abstract) [本文引用:1]
[23]	匡猛, 杨伟华, 许红霞, 王延琴, 周大云, 冯新爱, 王俊芳. 分子标记技术在棉花品种鉴定上的研究进展. 棉花学报, 2009, 21: 330-334 Kuang M, Yang W H, Xu H X, Wang Y Q, Zhou D Y, Feng X A, Wang J F. Research progress of molecular marker technology applied in cotton variety identification. Cotton Sci, 2009, 21: 330-334 (in Chinese with English abstract) [本文引用:1] [CJCR: 1.462]
[24]	Union for the Protection of New Varieties of Plants. Guidelines for Molecular Marker Selection and Database Construction//BMT Guidelines (Proj. 3). Geneva: UPOV, 2005. p326 [本文引用:1]
[25]	陆徐忠, 马琳, 倪金龙, 张小娟, 杨剑波. 基于DNA 指纹建立农作物品种身份证的方法探析. 中国农学通报, 2014, 30(9): 248-252 Lu X Z, Ma L, Ni J L, Zhang X J, Yang J B. Perspectives on the establishment of a crop identity system based on DNA fingerprinting. Chin Agric Sci Bull, 2014, 30(9): 248-252 (in Chinese with English abstract) [本文引用:1]
[26]	Coryell V H, Jessen H, Schupp J M, Webb D, Keim P. Allele- specific hybridzation markers for soybean. Theor Appl Genet, 1999, 98: 690-696 [本文引用:1] [JCR: 3.658]
[27]	曲鲁江, 李显耀, 杜志强, 张龙超, 杨宁. 微卫星PCR产物变性与非变性银染检测PAGE方法的比较. 遗传, 2004, 26: 522-524 Qu L J, Li X Y, Du Z Q, Zhang L C, Yang N. The comparision of denatured non-denatured PAGE-silver-sataining detection of PCR product of microsatellites. Hereditas (Beijing), 2004, 26: 522-524 (in Chinese with English abstract) [本文引用:1]
[28]	艾先涛, 梁亚军, 沙红, 王俊铎, 郑巨云, 吐尔逊江, 多力坤, 李雪源, 华金平. 新疆自育陆地棉品种SSR遗传多样性分析. 作物学报, 2014, 40: 369−379 Ai X T, Liang Y J, Sha H, Wang J D, Zheng J Y, Tu’er xunjiang, E X J, Duo L K, Li X Y, Hua J P. Genetic diversity analysis on local upland cotton cultivars in Xinjiang based. Acta Agron Sin, 2014, 40: 369−379 (in Chinese with English abstract) [本文引用:2] [CJCR: 1.667]
[29]	Liu D Q, Guo X P, Lin Z X, Nie Y C, Zhang X L. Genetic diversity of Asian cotton (Gossgpium arborreum L. ) in China evaluated by microsatellite analysis. Genet Resour Crop Evol, 2006, 53: 1145-1152 [本文引用:1]
[30]	Ni X Y, Lei W X, Zhao J Y. Establishment of molecular identity ID Brassica napus varieties using SSR markers. Cruciferous oil crops and sustainable development. In: The Paper Abstract of 12th International Conference on Rapeseed. Monmouth Junction: Scientific American Publishing Company, 2007. p166 [本文引用:1]
[31]	高运来, 朱荣胜, 刘春燕, 李文福, 蒋洪蔚, 李灿东, 姚丙晨, 胡国华, 陈庆山. 黑龙江部分大豆品种分子ID的构建. 作物学报, 2009, 35: 211-218 Gao Y L, Zhu R S, Liu C Y, Li W F, Jiang H W, Li C D, Yao B C, Hu G H, Chen Q S. Establishment of molecular ID in soybean varieties in Heilongjiang, China. Acta Agron Sin, 2009, 35: 211-218 (in Chinese with English abstract) [本文引用:1] [CJCR: 1.667]
[32]	宋海斌, 崔喜波, 马鸿艳, 朱子成, 栾非时. 基于SSR标记的甜瓜品种(系)DNA 指纹图谱库的构建. 中国农业科学, 2012, 45: 2676-2689 Song H B, Cui X B, Ma H Y, Zhu Z C, Luan F S. Construction of DNA fingerprint database based on SSR marker for varieties (lines) of Cucumis melo L. Sci Agric Sin, 2012, 45: 2676-2689 (in Chinese with English abstract) [本文引用:1] [CJCR: 1.889]
[33]	陆徐忠, 倪金龙, 李莉, 汪秀峰, 马卉, 张小娟, 杨剑波. 利用SSR分子指纹和商品信息构建水稻品种身份证. 作物学报, 2014, 40: 1-9 Lu X Z, Ni J L, Li L, Wang X F, Ma H, Zhang X J, Yang J B. Construction of rice variety based on ID used SSR fingerprint and commodity information. Acta Agron Sin, 2014, 40: 1-9 (in Chinese with English abstract) [本文引用:1] [CJCR: 1.667]

2012

0.0

... 随着转基因技术在棉花育种中的应用, 在原品种基础上改良少数或单个性状的衍生品种增多, 完全依据形态性状进行棉花品种的辨别越来越困难^[1] ...

2012

0.0

1.462

张玉翠, 杨伟华, 匡猛, 许红霞, 王延琴, 周大云, 冯新爱, 苏畅. 32个棉花主栽品种DNA指纹图谱构建及遗传多样性分析. 棉花学报, 2012, 24: 120-126

Zhang

Y C

, Yang

W H

, Kuang

, Xu

H X

, Wang

Y Q

, Zhou

D Y

, Feng

X A

, Su

Construction of DNA fingerprinting and analysis of genetic diversity with SSR markers for 32 leading cotton varieties. Cotton Sci, 2012, 24: 120-126 (in Chinese with English abstract)

Thirty-two leading cotton cultivars were used as materials in the construction of DNA fingerprinting and analysis of genetic diversity with simple sequence repeat(SSR) markers. Forty primer pairs with high polymorphisms, good repeatability, and even distribution on the 26 chromosomes were screened out from 95 candidates. A total of 161 polymorphic genotypes were detected among the 32 cultivars using the 40 SSR primer pairs, with an average of 4.025. The polymorphism information content ranged from 0.2989 to 0.7585, with an average of 0.5407 per primer. Eleven primer pairs produced specific genotype bands on 13 cultivars. These 32 cotton cultivars could be absolutely identified by just 5 primer pairs. NTSYSpc v2.10 software was used to analyze genetic diversity of the cultivars. The results indicated that the cultivars from the Yangtze River valley had the highest polymorphic level, followed by those from the Yellow River valley, with the level of cultivars from the Xinjiang region being the lowest. The heritability of conventional cultivars was narrower than the hybrids.

Cotton Research Institute of CAAS / State Key Laboratory of Cotton Biology, Anyang, Henan 455000, China

以我国2010年32个主栽品种为材料，利用SSR标记进行DNA指纹图谱的构建和遗传多样性分析。从95对SSR引物中，挑选出多态性高、稳定性好、均匀分布在棉花26条染色体上的40对引物，共扩增出161种多态性基因型，平均每对引物扩增出4.025种。引物多态信息量（PIC）0.2989～0.7585，平均为0.5407。11对引物在13个品种上有特征带型，最少利用5对引物就可以将32份材料完全区分开。利用NTSYS-pc V2.10软件聚类分析表明：长江流域棉区品种间遗传差异最大，黄河流域棉区次之，新疆棉区最小；常规种遗传基础较杂交种窄。

... 传统的棉花品种真实性和纯度是通过田间种植的方法鉴定的, 耗时长、成本高、时效性差, 易受环境因素的影响^[2] ...

... 与其他分子标记相比, 以微卫星序列为基础的SSR标记在DNA指纹鉴定上显示了独特的优越性, SSR 标记数量丰富, 覆盖整个基因组, 揭示的多态性高, 以孟德尔方式遗传, 呈共显性^[2] ...

... 张玉翠等^[2]用32个主栽品种, 40对引物, 共扩增出161种多态性基因型, 引物多态信息量(PIC)为0 ...

... 从附表看出, 新陆早9号、新陆早16、新陆早25、新陆早31和新陆早39的亲本均来源于贝尔斯诺系统, 因此B类品种主要亲本来源有贝尔斯诺、爱字棉系统、中棉所12、中棉所17、辽棉系列, 此系列选育品种数量最多、棉花纤维品质较优, 说明聚类结果和品种根据实际系谱来源分类是一致的, 这与张玉翠等^[2]的研究结果一致, 即遗传相似系数聚类分析结果基本上反映了品种之间的亲缘关系 ...

2012

1.643

0.0

... 近年来, 随着分子生物学的不断发展, 分子标记技术因不受环境条件的影响且多态性标记可反映DNA水平上生物个体间的遗传差异而被广泛应用, 其中SSR标记被认为是品种鉴定较为理想的标记之一^[3] ...

1996

0.0

... 目前, 分子标记技术广泛应用在棉花品种指纹图谱构建和遗传多样性分析中, 例如, 郭旺珍等^[4]最早在1996年利用RAPD技术对中国9个棉花主栽品种的指纹图谱进行了研究 ...

1995

1.668

0.0

... Multani等^[5]利用RAPD标记构建了澳大利亚13个陆地棉品种和1个海岛棉品种的指纹图谱 ...

1997

3.658

0.0

Iqbal

M J

, Aziz

, Saeed

N A

, Zafar

, Malik

K A.

Genetic diversity evaluation of some elite cotton varieties by RAPD analysis. Theor Appl Genet, 1997, 94: 139-144

1.National Institute for Biotechnology and Genetic Engineering (NIBGE), Jhang Road, P.O. Box 577, Faisalabad, Pakistan

Random amplified polymorphic DNA (RAPD) analysis was used to evaluate the genetic diversity of elite commercial cotton varieties. Twenty two varieties belonging to Gossypium hirsutum L. and one to G. arboreum L. were analyzed with 50 random decamer primers using the polymerase chain reaction (PCR). Forty nine primers detected polymorphism in all 23 cotton varieties, while one produced monomorphic amplification profiles. A total of 349 bands were amplified, 89.1% of which were polymorphic. Cluster analysis by the unweighted pair group method of arithmetic means (UPGMA) showed that 17 varieties can be placed in two groups with a similarity ranging from 81.51% to 93.41%. G. hirsutum L. varieties S-12, V3 and MNH-93 showed a similarity of 78.12, 74.46 and 69.56% respectively with rest of the varieties. One variety, CIM-1100, showed 57.02% similarity and was quite distinct. The diploid cotton G. arboreum L. var. Ravi was also very distinct from rest of its tetraploid counterparts and showed only 55.7% similarity. The analysis revealed that the intervarietal genetic relationships of several varieties is related to their center of origin. As expected, most of the varieties have a narrow genetic base. The results obtained can be used for the selection of possible parents to generate a mapping population. The results also reveal the genetic relationship of elite commercial cotton varieties with some standard “Coker” varieties and the diploid G. arboreum L. var. Ravi (old world cotton).

... Iqbal等^[6]对22个陆地棉品种和1个亚洲棉品种进行RAPD分析, 认为陆地棉品种的遗传基础很窄 ...

2001

0.0

1.462

... 武耀廷等^[7]利用SSR标记检测了30个陆地棉栽培品种和4个杂交种亲本的多态性, 并建立了杂交种的指纹图谱, 用于它们的分子鉴定和纯度检测 ...

2003

0.0

1.1628

马轩, 杜雄明, 孙君灵. 18个彩色棉品系的SSR指纹分析. 植物遗传资源学报, 2003, 4: 305-310

, Du

X M

, Sun

J L.

SSR fingerprinting analysis on 18 colored cotton lines. J Plant Genet Resourc, 2003, 4: 305-310 (in Chinese with English abstract)

本研究利用SSR技术建立了18个彩色棉品系的DNA指纹图谱,在110对SSR引物中筛选到10对扩增效果较好的引物,应用其中的4对构建了18个彩色棉的分子检索模式图.同时用这10对引物对18个彩色棉进行了聚类分析,对它们的亲缘关系作了初步探讨.

... 马轩等^[8]利用SSR技术建立18个彩色棉品系的DNA指纹图谱 ...

2007

0.0

0.951

姜伟, 李雪源, 何觉民, 陆建农. 新疆8个棉花品种(系)的指纹图谱分析. 华北农学报, 2007, 22: 89-92

Jiang

, Li

X Y

, He

J M

, Lu

J N.

Fingerprinting analysis for 8 cultivars (lines) of Xinjiang. Acta Agric Boreali-Sin, 2007, 22: 89-92 (in Chinese with English abstract)

利用ISSR对8个新疆品种(系)进行指纹图谱构建.通过基因组多态性分析,从60条引物中筛选到11条扩增效果好的引物,用其中2条引物UBC809,UBC841建立8个品种的指纹图谱,统计品种鉴定的置信概率达到1/8388608.结果显示,ISSR分子标记技术对棉花品种的指纹图谱构建具有高效性和准确性.

... 姜伟等^[9]利用ISSR标记, 对8个新疆品种(系) 构建指纹图谱 ...

2008

0.0

1.667

李武, 倪薇, 林忠旭, 张献龙. 海岛棉遗传多样性的SRAP标记分析. 作物学报, 2008, 34: 893−898

, Ni

, Lin

Z X

, Zhang

X L.

Genetic diversity analysis of sea-island cotton cultivars using SRAP markers. Acta Agron Sin, 2008, 34: 893−898 (in Chinese with English abstract)

Sea-island cotton (Gossypium barbadense) is important due to its fine fiber quality. However, genetic diversity analysis of this specie is still poorly documented, which is important for in situ and ex situ germplasm conservation and useful for breeding programs. Using 132 out of 238 SRAP markers, genetic diversity of 56 sea-island cotton cultivars including 20 introduced from other countries was assessed, 419 polymorphism loci were detected. The number of polymorphic loci per primer ranged from 2 to 9 with an average of 3.17. Jaccard’s genetic similarity coefficients were calculated and dendrograms were constructed by the unweighted pair group method of arithmetic average (UPGMA) using the NTSYS-pc version 2.10. The most of the accessions with relative relationship in their pedigrees were clustered into the same group. The genetic similarity coefficient of 56 accessions ranged from 0.312 to 0.876, with an average of 0.497, indicating that there was high level of genetic variation in the G. barbadense collections. The average genetic similarity coefficients of the domestic accessions in the first, second and third breed-ing periods were 0.501, 0.507, and 0.548, respectively, which implied that the genomic difference among the modern accessions has been gradually decreasing compared to that among the early accessions. These results suggest an overview of the genetic di-versity of the sea-island cotton accessions, and provide a reference for breeders to develop new cultivars efficiently.

1 National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan 430070, Hubei; 2 Economic Crop Research Institute, Henan Academy of Agricultural Sciences, Zhengzhou 450002, Henan, China

利用SRAP标记, 选用132对带型清晰的多态性引物, 对我国引入海岛棉以来培育的36个国内品种及20个国外品种进行遗传多样性分析。共检测到419个多态性位点, 每组合的多态性条带数从2~9不等, 平均为3.17。利用NTSYS-pc 2.10e 软件采用Jaccard’s相似系数和UPGMA方法进行聚类分析。结果表明, 大部分具有亲缘关系的品种聚在同类中, 说明其结果与系谱具有一定的相符性; 56个品种的平均遗传相似系数为0.497, 变化范围在0.312~0.876之间, 说明我国海岛棉品种在分子水平上存在较大差异; 我国3个育种时期育成品种的平均相似系数依次为0.501、0.507和0.548, 表明我国现在育成的品种相对于早期品种遗传多样性在逐渐降低。这些结果为我国海岛棉育种提供了有益的参考。

... 李武等^[10]利用SRAP标记, 对中国引入海岛棉以来培育的36个国内品种及20个国外品种进行了遗传多样性分析 ...

2010

0.0

... 潘兆娥等^[11]构建中棉所48的SSR数字指纹图谱 ...

2011

0.0

0.4744

... 薛艳等^[12]建立新疆早熟棉品种SSR分子标记体系 ...

... 本试验的聚丙烯凝胶电泳采用变性胶, 而薛艳等^[12]研究42份新疆陆地早熟棉指纹图谱构建与品种鉴别时采用9%非变性聚丙烯酰胺凝胶电泳分离方法, 曲鲁江等^[27]报道的非变性凝胶中条带过于复杂, 出现的非特异性条带极大地影响了分析带形的准确性, 使试验结果的误差加大, 变性胶在利用银染检测微卫星产物时条带清晰, 带型分析准确性更高, 结果更加可靠 ...

2011

0.0

1.462

李成奇, 王晓芸, 张晓飞, 王清连. 百棉系列棉花自交系品种(系)SSR指纹图谱构建. 棉花学报, 2011, 23: 228-234

Ch Q

, Wang

X Y

, Zhang

X F

, Wang

Q L.

SSR fingerprinting establishment of baimian series cotton inbred varieties (lines). Cotton Sci, 2011, 23: 228-234

SSR markers were used to analyze the DNA fingerprinting of Baimian series cotton inbred varieties(lines) with 32 cotton background materials mainly rooted in Deltapine, Stoneville, Foster, and Uganda. 20 SSR primers selected from 2503 SSR primers, with high polymorphisms and good repeatability, were regarded as core primers in the study. These primers belonged to 15 chromosomes of cotton and 116 alleles were detected by them. The average alleles, polymorphic information content and marker index coefficient of 20 SSR primers were respectively 5.8, 0.718 and 4.384, indicating that they had preferable polymorphism. Primer combinations NAU1028/NAU5104, NAU4903/NAU3110/NAU1043, NAU0905/NAU1255, NAU3839/ NAU4024, NAU5104/NAU4024, NAU2121/NAU5104 and NAU1043/NAU3254/NAU4024 could be used to distinguish Baimian 1, Baimian 2, Baimian 13, Baimian 5, Baimian 011, Baimian 19, and Baimian 985 from other materials, respectively. SSR digital fingerprinting code of Baimian series cotton inbred varieties(lines) was also established. The experimental materials and the core primers during the fingerprinting establishment of cotton were analyzed and discussed.

Henan Institute of Science and Technology, Key Discipline Open Laboratory on Crop Molecular Breeding, Universities of Henan Province / Cotton Research Institute, Xinxiang, Henan 453003, China

利用20对SSR核心引物构建了百棉系列棉花自交系品种（系）的DNA指纹图谱。20对引物分属棉花15条染色体，共检测到116个等位基因，平均5.8个；PIC值和MI值分别平均为0.718和4.384。引物组合NAU1028/NAU5104、NAU4903/NAU3110/NAU1043、NAU0905/NAU1255、NAU3839/NAU4024、NAU5104/NA U4024、NAU2121/NAU5104和NAU1043/NAU3254/NAU4024可分别将百棉1号、百棉2号、百棉13号、百棉5号、百棉011、百棉19号和百棉985与其他所有材料区分开。构建了百棉系列棉花自交系品种（系）的SSR数字指纹代码。对棉花指纹图谱构建的供试材料和核心引物进行了分析和讨论。

... 李成奇等^[13]利用20对SSR核心引物构建了百棉系列棉花自交系品种(系)的DNA指纹图谱 ...

2011

0.0

1.462

孙宁, 杨付新, 付小琼, 王秀玲, 刘逢举, 李亚丽, 汤磊鹏. 国家棉花区试新品种的SSR指纹图谱分析. 棉花学报, 2011, 23: 279-283

Sun

, Yang

F X

, Fu

X Q

, Wang

X L

, Liu

F J

, Li

Y L

, Tang

L P.

SSR marker fingerprinting of cotton varieties in national regional trails. Cotton Sci, 2011, 23: 279-283 (in Chinese with English abstract)

In this study, 57 cotton cultivars in 2009 national regional trails were used as plant materials for DNA fingerprinting. Among 1800 pairs of primers, 22 pairs with steady polymorphism were selected. Totally, 145 bands were observed, and 106 (72.7%) were polymorphic, with an average of 4.8 bands per primer. The mean PIC value of these markers was 0.661, and a DNA fingerprint database for the varieties tested was developed. UPGMA cluster analysis of Genetic similarity showed that the observed 106 polymorphic markers are sufficient to separate all of the 57 varieties from one another at the maximum genetic similar coefficient of 0.98. Cultivars derived from the same district or bred by same institutes were clustered together in different extent, indicating their close genetic relationship.

Cotton Research Institute of CAAS/ Key Laboratory of Cotton Genetic Improvement, Ministry of Agriculture, Anyang, Henan 455000, China

以2009年度参加黄河流域国家棉花区试的57份参试品种为材料，运用筛选确定的22对多态性好、条带清晰的SSR引物对参试品种进行分子标记分析，共检测出145个等位位点，其中106个为多态性位点，平均每对引物4.8个，多态性比率平均达到72.7%，PIC平均值为0.661。利用106个多态性片段构建了这些品种的SSR指纹图谱。遗传聚类结果显示，在相似系数为0.98时，57个区试品种完全区分开；同一地区和同一单位育成的品种在不同程度上聚在一类，说明它们的遗传关系较近。

... 孙宁等^[14]以2009年度参加黄河流域国家棉花区试的57份参试品种为材料进行指纹图谱分析 ...

2011

0.0

1.889

匡猛, 杨伟华, 许红霞, 王延琴, 周大云, 冯新爱. 中国棉花主栽品种DNA指纹图谱构建及SSR标记遗传多样性分析. 中国农业科学, 2011, 44: 20-27

Kuang

, Yang

W H

, Xu

H X

, Wang

Y Q

, Zhou

D Y

, Feng

X A.

Construction of DNA fingerprinting and analysis of genetic diversity with SSR markers for cotton major cultivars in China. Sci Agric Sin, 2011, 44: 20-27 (in Chinese with English abstract)

【Objective】The aim of this study was to construct a DNA fingerprinting database of 32 cotton major cultivars from three main cotton regions (including eight main cotton production provinces), and the genetic diversity was analyzed based on simple sequence repeats (SSR).【Method】Thirty six evenly distributed SSR primer pairs with high polymorphisms and good repeatability were successfully screened out from 214 candidates to construct the fingerprinting database.【Result】Among the 32 varieties, 36 primer pairs had 142 polymorphic genotypes, and 3.94 genotypes were detected by each SSR primer pair on an average with the range from 2 to11. Ten cultivars had specific genotypes by 9 primer pairs. Thirty two major cultivars could be identified by 5 primer combinations at least. The genetic diversity analyzed by the software of NTSYS-pc V 2.10 indicated that Yangtze River valley cotton region had the highest polymorphic level, followed by Xinjiang, and the lowest was the varieties from the Yellow River valley. The genetic basis of conventional cultivars was narrower than that of hybrids. 【Conclusion】 Core primer combination method is better than distinct band method for the construction of DNA fingerprinting of cotton major cultivars in China. The genetic relationship of the cotton cultivars derived from the same district is similar to a certain extent.

(中国农业科学院棉花研究所/农业部棉花遗传改良重点开放实验室)

【目的】采用SSR标记构建2008年中国3大棉区8个棉花主产省份32份棉花主栽品种的DNA指纹图谱并进行遗传多样性分析。【方法】从214对候选引物中筛选出36对多态性高、稳定性好且在染色体上分布均匀的引物作为核心引物,构建棉花主栽品种DNA指纹图谱。【结果】36对SSR引物在32份材料中共扩增出142种多态性基因型,每对引物的基因型从2—11种不等,平均每对引物扩增出3.94种基因型。9对引物在10个品种上具有特征带型。最少采用5对引物进行组合鉴定即可将32个棉花品种完全区分开。NTSYS-pc V 2.10软件分析表明,长江流域棉区品种间遗传差异最大,新疆棉区次之,黄河流域棉区最小。常规品种的遗传基础窄于杂交陆地棉。【结论】核心引物组合法相比特征谱带法更适用于构建中国棉花主栽品种DNA指纹图谱,品种间的亲缘关系与地理来源有一定的相关性。

... 匡猛等^[15]采用SSR标记构建2008年中国3大棉区8个棉花主产省份32份棉花主栽品种的DNA指纹图谱并进行遗传多样性分析 ...

2012

0.0

1.667

赵亮, 蔡彩平, 梅鸿献, 郭旺珍. 用于区别不同棉花品种基因组特征的微卫星位点筛选. 作物学报, 2012, 38: 1810-1817

Zhao

, Cai

C P

, Mei

H X

, Guo

W Z.

Screening of microsatellite loci for identifying genome barcoding of cotton cultivars. Acta Agron Sin, 2011, 44: 20-27 (in Chinese with English abstract)

Microsatellites or simple sequence repeats (SSRs) loci with high information content, reproducibility and locus specificity can be used effectively to construct DNA barcode in plant. In this study, 12 cotton cultivars derived from different germplasm pedigrees and planted widely in three ecological cotton growing areas were selected to screen the SSR loci with high polymorphism. Based on our newly updated high-density genetic linkage map from G. hirsutum × G. barbadense BC1 mapping population, we selected 51 primer pairs with clear amplification products and high polymorphism from 376 genome-wide SSR primer pairs. In total, 155 polymorphic loci in 12 cultivars were produced by using the selected 51 primer pairs. Alleles ranging from two to seven with an average of 3.04 were detected by each SSR primer pair. Of them, 26 SSR primer pairs, which amplified SSR loci were tagged on corresponding 26 chromosomes in cultivated tetraploid cotton species, were recommended as first selected primer pairs to establish DNA barcode of cotton cultivars, and other 25 SSR primer pairs could be used as candidate primer pairs. DNA barcodes of 12 cultivars were further constructed effectively using these microsatellite loci with high polymorphism. In the future, these microsatellite loci can be used to construct DNA barcode for a large number of cotton varieties, and applied effectively in the identification of genuineness and purity in cotton cultivar seeds.

State Key Laboratory of Crop Genetics and Germplasm Enhancement, Hybrid Cotton R&D Engineering Research Center, Ministry of Education, Nanjing Agricultural University, Nanjing 210095, China

保守性强、重复性好、多态性高的微卫星位点可被有效用于构建作物DNA条形码。选取目前生产上主要推广种植、代表不同来源系统的12个棉花品种作为微卫星位点筛选材料，参考我室构建的四倍体栽培棉种种间高密度遗传图谱信息，从376对覆盖全基因组的SSR引物中，筛选出51对引物可扩增出带型清晰且多态性高的微卫星位点。这些引物在12个供试品种中共产生155个等位位点，每对引物揭示的等位基因位点在2~7之间，平均值为3.04。参照微卫星位点的染色体定位和多态信息，在每条染色体上选择一个多态性相对高的SSR位点，其相应的26对SSR引物被推荐为构建棉花品种DNA条形码的一套首选引物，并初步应用于12个品种的DNA条形码编制。其余25对引物作为候选引物。使用该套引物扩增出的微卫星位点可用于大量棉花品种DNA条形码构建，为棉花品种真实性和纯度的分子鉴定奠定基础。

... 赵亮等^[16]利用26对SSR引物构建12个品种DNA条形码编制 ...

... 与赵亮^[16]等的研究具有明显差异, 本试验选用新疆2013年前审定的早熟陆地棉品种全套完整材料, 75对核心标记均匀分布于棉花26条染色体且较高多态性, 结合特征引物和组合引物方法构建品种指纹 ...

2007

0.0

0.916

程保山, 万志兵, 洪德林. 35个粳稻品种SSR指纹图谱的构建及遗传相似性分析. 南京农业大学学报, 2007, 30(3): 1-8

Cheng

B S

, Wan

Z B

, Hong

D L.

Establishment of SSR fingerprint map and analysis of genetic similarity among 35 varieties in japonica rice. J Nanjing Agric Univ, 2007, 30(3): 1-8 (in Chinese with English abstract)

用68对SSR引物扩增了35个粳稻品种(系)的基因组DNA,结果有46对引物在35个品种间具有稳定多态性.有6个品种可用单一特异的SSR标记加以识别,其余29个品种则需要不同的SSR指纹组合才能识别.用12对核心引物构建的SSR指纹图谱能将35个粳稻品种逐一区别开来.35个粳稻品种间遗传相似系数的变异范围为0.27～0.98.采用类平均法进行聚类分析,在相似系数0.82处,可将35个粳稻品种分为4类.聚类分析结果基本上反映了依据系谱分析的品种间亲缘关系.

... 近年来, 基于SSR标记构建了水稻^[17]、小麦^[18]、玉米^[19]、烟草^[20]、大豆^[21]等主要农作物的DNA指纹图谱数据库 ...

2006

0.0

1.667

李根英, Dreisigacker

, Warburton

M L

, 夏先春, 何中虎, 孙其信. 小麦指纹图谱数据库的建立及SSR分子标记试剂盒的研发. 作物学报, 2006, 32: 1771-1778

G Y

, Dreisigacker

, Warburton

M L

, Xia

X C

, He

Z H

, Sun

Q X.

Development of a fingerprinting database and assembling an SSR reference kit for genetic diversity analysis of wheat. Acta Agron Sin, 2006, 32: 1771-1778 (in Chinese with English abstract)

Understanding of the current and expanded genetic diversity is very important for raising the yield of wheat. Genetic diversity based on molecular markers has been studied in plants for over thirty decades. SSR is the currently most popular marker system in wheat. In order to utilize the diversity held in NARS (National Agricultural Research Station) and CGIAR (Consultative Group on International Agricultural Research) germplasm collections, one of the GCP’s (Generation Challenge Program) premier capacity building activities is to build databases that contain traditional and molecular data on germplasm so that scientists all over the world can access information with relevance to their region on traits, genes, and sequences. In the present study, a fingerprinting database was established containing 134 SSR primers and 2 457 wheat genotypes with the data from CIMMYT and three collaborators: ICARDA, Agropolis, and CAAS. On the base of the database, a SSR reference kit for wheat genetic diversity analysis was developed, which will facilitate the use of this data in new projects and cross-laboratory comparisons. In total, 46 SSR primers with comparatively high polymorphism were selected as the reference markers to constitute the standard allele kit, 334 genotypes fingerprinted within the GCP tier 1 project “Genotyping a composite germplasm set in wheat” were chosen to represent the SSR allele kit consisting of 794 alleles amplified by 46 SSR markers. Genotypes were originally selected and DNA extracted by CIMMYT and 3 additional collaborators: ICARDA, Agropolis, and CAAS. These 334 genotypes, when taken as a group, amplify every allele seen for the wheat genotypes for all the 46 SSRs. The Genotypes were originally amplified with 26 SSRs at CIMMYT, with 8 SSRs at INRA, France, and with 12 SSRs at CAAS. Genotypes were amplified with all SSRs at CIMMYT again to confirm the results. For reamplification at CIMMYT, forward primers were labeled at the 5’ end with either one of three phosphoramidite fluorescent dyes 6-carboxyflouresein (6-Fam), tetrachloro-6-carboxyflouresein (Tet) or hexachloro-6- carboxyflouresein (Hex). PCR-reactions were carried out in an MJ-Research thermocycler model PTC225. Amplification products were separated on an ABI-Prism SequencerTM377 using 4.5% polyacrylamide denaturing gels. Fragment sizes were calculated semi-automatically with the computer software GeneScan 3.1 by comparing to fragments of an internal size standard (GeneScan 350 or 500) labeled with N,N,N,N, -tetramethyl-6-carboxyrhodamine (Tamra). GeneScan fragments were assigned to alleles using the category function of the software Genotyper 2.1. The two genotypes, Opata and Synthetic were run in each gel as controls.
Finally, a SSR reference kit was assembled which includes 46 pairs of SSR primers, protocols (PCR condition, detection of polymorphisms, etc), description of polymorphism in the reference samples, reference DNA samples for a complete range of repeatable 592 alleles, methods for comparing new to preexisting data, and classification of various genotypes. The database and the reference kit will provide a powerful tool for genetic diversity studies of wheat germplasm worldwide.

1 College of Agronomy and Biotechnology, China Agricultural University, Beijing 100094, China; 2 Institute of Crop Sciences/National Wheat Improvement Center/ National Key Facility for Crop Gene Resources and Genetic Improvement, Chinese Academy of Agricultural Sciences, Beijing 100081, China; 3International Maize and Wheat Improvement Center (CIMMYT), Apdo. Postal 6-641, 06600, Mexico D.F., Mexico; 4 CIMMYT China Office, Beijing 100081, China; 5Crop Research Institute, Shandong Academy of Agricultural Sciences, Jinan 250100, China

本研究以国际玉米小麦改良中心（CIMMYT）、国际干旱地区农业研究中心（ICARDA）、法国Agropolis研究所和中国农业科学院作物科学研究所提供的数据为基础，建立了包括134个SSR引物、2457个普通小麦基因型的指纹图谱数据库。利用荧光标记法的分析结果，用代表性基因型在某一位点的扩增片段作为银染法的读带依据，开发了SSR分子标记试剂盒，包括46对SSR引物及其PCR反应程序、代表性基因型的DNA样品、592个等位变异的代表性基因型清单及试剂盒使用说明等。该数据库的建立和试剂盒的开发，为利用SSR分子标记技术进行小麦种质遗传多样性研究，实现小麦遗传资源和信息资源全球共享提供了重要工具和技术平台。

... 近年来, 基于SSR标记构建了水稻^[17]、小麦^[18]、玉米^[19]、烟草^[20]、大豆^[21]等主要农作物的DNA指纹图谱数据库 ...

2003

0.0

0.965

... 近年来, 基于SSR标记构建了水稻^[17]、小麦^[18]、玉米^[19]、烟草^[20]、大豆^[21]等主要农作物的DNA指纹图谱数据库 ...

2011

0.0

1.2747

徐军, 刘艳华, 任民, 牟建民, 张兴伟, 陈雅琼, 王志德. 普通烟草种质资源的SSR标记与指纹图谱分析. 中国烟草科学, 2011, 32(2): 62-65

, Liu

Y H

, Ren

, Mou

J M

, Zhang

X W

, Chen

Y Q

, Wang

Z D.

SSR fingerprint map analysis of tobacco germplasms. Chin Tob Sci, 2011, 32(2): 62-65 (in Chinese with English abstract)

利用SSR标记构建了烟草核心种质中80份普通烟草种质的指纹图谱.从286对SSR引物中筛选出8对多态性较好的引物,对这些烟草种质进行分析,共检测到85个多态性位点.研究表明,这8对多态性SSR引物可以将80份烟草种质完全区分开,每份种质都有各自独特的指纹图谱,表明SSR标记完全适用于普通烟草种质鉴定和指纹图谱构建工作.

... 近年来, 基于SSR标记构建了水稻^[17]、小麦^[18]、玉米^[19]、烟草^[20]、大豆^[21]等主要农作物的DNA指纹图谱数据库 ...

2001

0.0

1.889

田清震, 盖钧镒, 喻德跃, 吕慧能, 贾继增. 我国野生大豆与栽培大豆AFLP指纹图谱研究. 中国农业科学, 2001, 34: 465-468

Tian

Q Z

, Gai

J Y

, Yu

D Y

, Lü H N, Jia J Z. AFLP fingerprint analysis of G. soja and G. max in China. Sci Agric Sin, 2001, 34: 465-468 (in Chinese with English abstract)

浙江大学原子核农业科学研究所

用 Envirologix Cry1Ab/Cry1Ac平板试剂盒检测了两个转 cry1Ab的水稻品系克螟稻 1号 ( KMD1)、克螟稻 2号 ( KMD2 ) ,及未转化的对照品种秀水 11米粒中的 Bt杀虫蛋白含量。结果表明 ,该试剂盒标样制作的标准曲线相关系数为 0 .985～ 0 .998,在 0 .0 5或 0 .0 1水平上显著。同批次试剂盒的不同试验及不同批次试剂盒的测定值均差异不显著。最低检测剂量达 0 .5 ng/g,每次测试时间比 Western dot blotting方法缩短约 2 h。试验表明 ,该试剂盒是定量检测转基因水稻 Bt毒蛋白表达的一种理想产品

... 近年来, 基于SSR标记构建了水稻^[17]、小麦^[18]、玉米^[19]、烟草^[20]、大豆^[21]等主要农作物的DNA指纹图谱数据库 ...

2011

0.0

... 2 DNA提取及SSR检测参照白静的棉花总DNA提取方法、SSR-PCR反应体系、PCR扩增程序、PAGE凝胶电泳及显色方法^[22] ...

2009

0.0

1.462

匡猛, 杨伟华, 许红霞, 王延琴, 周大云, 冯新爱, 王俊芳. 分子标记技术在棉花品种鉴定上的研究进展. 棉花学报, 2009, 21: 330-334

Kuang

, Yang

W H

, Xu

H X

, Wang

Y Q

, Zhou

D Y

, Feng

X A

, Wang

J F.

Research progress of molecular marker technology applied in cotton variety identification. Cotton Sci, 2009, 21: 330-334 (in Chinese with English abstract)

There are three phases of DNA fingerprint technology based on molecular marker：the first is RFLP based on Southern blot，the second are several markers based on PCR and the third is SNP（single nucleotide polymorphism）.Research and application of variety identification technology was discussed，four main molecular marker technologies applied in cotton variety identification were introduced in detail as follow:RFLP,RAPD,AFLP and SSR.Technology principle,research and application status,advantage and disadvantage about each molecular marker applied in cotton variety identification were expatiated.As compared，the viewpoint was put forward:SSR marker was the most appropriate molecular marker in cotton variety identification,and the necessity of construction of Chinese cotton DNA fingerprint database based on SSR marker technology was analyzed.

Cotton Research Institute,CAAS; Supervision,Inspection and Test Center of Cotton Quality,Ministry of Agriculture,Anyang, Henan 455000，China

DNA指纹技术的发展大体分为三个阶段：第一代的分子标记是以Southern杂交为基础的RFLP，第二代分子标记是以PCR为基础的各种DNA指纹标记，第三代分子标记是以单核苷酸多态性为基础的SNP。本文概述了品种鉴定技术的研究与应用进展，重点介绍了用于棉花品种鉴定的四种主要的分子标记技术：RFLP、RAPD、AFLP和SSR，对每一种分子标记的技术原理、在棉花品种鉴定上的应用研究状况及存在的优缺点进行了具体阐述。通过比较分析，提出了SSR标记适用于棉花品种鉴定的独特优越性。并对基于SSR分子标记技术构建我国棉花品种DNA指纹库的重要意义与必要性进行了分析。

... DNA指纹技术的发展经历3个阶段, 第1代是以Southern杂交为基础的RFLP, 第2代是以PCR为基础的DNA指纹标记(如RAPD、AFLP、SSR、SCAR、ISSR等), 第3代是以单核苷酸多态性为基础的SNP^[23] ...

2005

0.0

... 近些年, 分子标记开始应用于各物种的鉴定, 国际植物品种权保护组织(UPOV)在BMT测试指南草案中已将构建DNA指纹数据库的标记确定为SSR和SNP^[24] ...

2014

0.0

陆徐忠, 马琳, 倪金龙, 张小娟, 杨剑波. 基于DNA 指纹建立农作物品种身份证的方法探析. 中国农学通报, 2014, 30(9): 248-252

X Z

, Ma

, Ni

J L

, Zhang

X J

, Yang

J B.

Perspectives on the establishment of a crop identity system based on DNA fingerprinting. Chin Agric Sci Bull, 2014, 30(9): 248-252 (in Chinese with English abstract)

Crop identity (ID) cards can greatly assist the scientific recognition and management for crop cultivars. This article reviews the necessity and feasibility of an ID card system, proposes a format for cultivar ID cards, and discusses the establishment of such a system, with the goal of a more rational and efficient process. Crop cultivar ID cards should include three components: (1) the 11-character commodity code contains basic merchandise information, including crop, cultivar, and breeding (or approval) region and time; (2) the DNA fingerprint code could vary from 20–40 characters, depending on crop type; and (3)a supplementary code describing cultivar-specific gene information using the letter "T" with the gene name to represent GMO crop, "S" with the gene name to represent MAS crop,and "M" with the gene name to represent mutant line, etc.The complete code would be about 50 characters long. Cultivar ID card for a given seed could be expressed as a barcode, a two-dimensional code, or an electronic security code. Cultivar ID card numbers encoded by this method would be highly accurate, versatile, and traceable and would facilitate scientific and standardized management of crop cultivars in China.

构建农作物品种身份证对科学识别和管理农作物品种具有十分积极的意义。本文论述了建立品种身份证的必要性和可行性，阐述了品种身份证设计的基本思路及建立品种身份证制度的问题。指出：农作物品种身份证信息应由3部分构成：一是品种的基本商品信息，包括作物及品种类别、品种选育（或审定）的区域和时间等；二是品种DNA的一般指纹信息；三是品种的特异基因信息。按上述内容将身份证号码设计为：第1部分称为商品码，预设11位阿拉伯数字（或字母）；第2部分称为指纹码，依据作物类型不同预设20-40位阿拉伯数字（或字母）；第3部分称为补充码（特异基因识别码），设计为字母“T”（转基因育种）、“S”（分子育种）和“M”（诱变育种）等，加上被导入、渗入或诱变的基因名称；总位数约50位。品种身份证可以条形码、二维码或电子芯片方式表达，用于农作物种子身份的标识等。采用本方法编码的品种身份证号码，具有准确性高、通用性强及可追溯性等优越性。

... 随着第2代测序技术的发展, 使品种基因组DNA测序和SNP全面分析成为可能, 从而大大提升了品种识别和鉴定精度, 但检测成本较高, 有时会因检测尺度的过度精细, 把品种内的差异误判成品种间的差异^[25] ...

1999

3.658

0.0

Coryell

V H

, Jessen

, Schupp

J M

, Webb

, Keim

Allele- specific hybridzation markers for soybean. Theor Appl Genet, 1999, 98: 690-696

A1.Department of Biological Sciences, Northern Arizona University, Flagstaff, AZ 86011-5640, USA Fax: +1-520-523-0639 E-mail: paul.keim@nau.edu, US A2.Soybean Research Department, Pioneer Hi-Bred International, Johnston, IA 50131, USA, US

Soybean (Glycine max) is one of the world’s most important crop plants due to extensive genetic improvements using traditional breeding approaches. Recently, marker-assisted selection has enhanced the ability of traditional breeding programs to improve soybeans. Most methods of assessing molecular markers involve electrophoretic techniques that constrain the ability to perform high-throughput analyses on breeding populations and germplasm. In order to develop a high-capacity system, we have developed allele-specific hybridization (ASH) markers for soybean. As one example, restriction fragment length polymorphism (RFLP) locus A519-1 (linkage group B) was converted into an ASH marker by (1) sequencing the pA519 cloned insert, (2) designing locus-specific PCR amplification primers, (3) comparative sequencing of A519-1 amplicons from important soybean ancestors, and (4) designing allele-specific oligonucleotide probes around single nucleotide polymorphisms (SNPs) among soybean genotypes. Two SNPs were identified within approximately 400 bp of the sequence. Allele-specific probes generated a 100-fold greater signal to target amplicons than to targets that differed by only a single nucleotide. The A519-1 ASH marker is shown to cosegregate with the A519-1 RFLP locus. In order to determine ASH usefulness, we genotyped 570 soybean lines from the Pioneer Hi-Bred soybean improvement using both A519-1 SNPs. Combined haplotype diversity (D) was 0.43 in this adapted germplasm set. These results demonstrate that ASH markers can allow for high-throughput screening of germplasm and breeding populations, greatly enhancing breeders’ capabilities to do marker-assisted selection.

... 因此, SNP高额的投入使其未能得到广泛应用^[26], 其中SSR标记技术比较成熟, 成为当前各个作物构建指纹数据库的首选技术 ...

2004

0.0

2014

0.0

1.667

艾先涛, 梁亚军, 沙红, 王俊铎, 郑巨云, 吐尔逊江, 多力坤, 李雪源, 华金平. 新疆自育陆地棉品种SSR遗传多样性分析. 作物学报, 2014, 40: 369−379

X T

, Liang

Y J

, Sha

, Wang

J D

, Zheng

J Y

, Tu’er

xunjiang

, E

X J

, Duo

L K

, Li

X Y

, Hua

J P.

Genetic diversity analysis on local upland cotton cultivars in Xinjiang based. Acta Agron Sin, 2014, 40: 369−379 (in Chinese with English abstract)

Xinjiang, the largest prior cotton production region in China, has made remarkable genetic improvement in upland cotton (Gossypium hirsutum L.). There has been a substantial raise in yield potential, cotton-planting area and total production in Xinjiang since 1970s. The new upland cotton cultivars have contributed a lot to yield increase, in which Xinjiang-bred upland cotton varieties play an important role in cotton production in both Northern and Southern Xinjiang. However, seldom research on genetic basis of genetic diversity and pedigree relationship has been evaluated in upland cotton varieties in Xinjiang so far. This study aimed to reveal the genetic diversity of 94 upland cotton varieties in Xinjiang by genome analysis using SSR markers. The results indicated that SSR molecular markers could be employed in revealing the genetic diversity. One hundred and fifty-three polymorphic lociwere detected by amplified with 54 pairs of primers with allelic variation of 2-6 per primer pair and the average of 2.93. The genotypic diversity (H’) spanned from 0.0439 to 0.7149, with the average of 0.4491. The PIC value ranged from 0.0430 to 0.6640, and the average was 0.3831. The coefficients of genetic similarity among 94 tested cultivars varied from 0.3846 to 0.9835. And more than 71.9% of similarity coefficients fell into the interval of 0.601–0.800, indicating that the genetic similarity among the cultivars was high, whereas the genetic diversity might not be abundant. According to UPGMA cluster analysis, 94 tested cultivars were divided into two clusters when the threshold valve was given as 0.63, showing the relatively simple genetic relationship among the upland cotton varieties in Xinjiang, and relatively low genetic diversity and narrow genetic basis. The results of molecular clustering on genetic diversity of upland cotton coincided in large degree with the pedigree relationship. In conclusion, there is no much difference on molecular level among Xinjiang-bred upland cotton varieties. Materials with much wider genetic backgrounds are suggested to be used to diversify the genetic basis of cotton varieties in Xinjiang.

1Xinjiang Agricultural University , Urumqi 830052, China; 2 Economic Crop Research Institute, Xinjiang Academy of Agricultural Sciences, Urumqi 830091, China; 3 Department of Plant Genetics and Breeding / Key Laboratory of Crop Heterosis and Utilization of Ministry of Education / Beijing Key Laboratory of Crop Genetic Improvement, China Agricultural University, Beijing 100193, China

利用SSR标记对94份新疆自育陆地棉品种的基因组进行分析，研究新疆陆地棉品种的遗传多样性。结果: 从分布于棉花全基因组的206对SSR标记中筛选出54对具有稳定多态性的引物, 共检测出153个多态性位点, 每对引物的等位变异为2~6个，平均为2.93个；基因型多样性(H′)变幅为0.0439–0.7149，平均为0.4491；引物多态信息含量(PIC)为0.0430–0.6640，平均为0.3831。表明SSR标记在品种间可以反映较丰富的遗传多样性信息。94份品种间成对遗传相似系数变幅为0.3846~0.9835, 71.9%的品种相似系数在0.601~0.800内，反映出新疆陆地棉品种间的遗传相似性相对较高。根据UPGMA 聚类分析，在阈值为0.63时，将94份品种划分为2个类群，说明新疆陆地棉品种间遗传关系相对简单，品种的遗传基础相对狭窄，品种遗传组分差异较小，总体上遗传多样性不够丰富；分子聚类结果与品种本身遗传系谱背景和演变趋势吻合度较高，符合品种的真实特性。研究证明，自育品种在分子水平上差异不大，需要努力拓宽品种选育的遗传基础。

... 艾先涛等^[28]用94份新疆陆地棉品种, 54对具有稳定多态性的引物, 共检测出153个多态性位点, 每对引物的等位变异为2~6个, 平均为2 ...

... 51个新陆早品种遗传相似系数矩阵和品种聚类表明, 新陆早棉花品种之间遗传多样性较狭窄, 这与艾先涛等^[28]的研究结果相似, 说明新疆陆地棉品种间遗传关系相对简单, 品种的遗传基础相对狭窄, 品种遗传组分差异较小, 总体上遗传多样性不够丰富 ...

2006

0.0

Liu

D Q

, Guo

X P

, Lin

Z X

, Nie

Y C

, Zhang

X L.

Genetic diversity of Asian cotton (Gossgpium arborreum L. ) in China evaluated by microsatellite analysis. Genet Resour Crop Evol, 2006, 53: 1145-1152

1.National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan, Hubei, 430070, China

Asian cotton (Gossypium arboreum L.) was once widely cultivated in China. It has also been a valuable source of genetic variation in modern cotton improvement. In this study, the genetic diversity of selected G. arboreum accessions collected from different regions of China was evaluated by microsatellite (simple sequence repeats, SSRs) analysis. Of the 358 microsatellite markers analyzed, 74 primer pairs detected 165 polymorphic DNA fragments among 39 G. arboreum accessions examined. Twelve accessions could be fingerprinted with one or more SSR markers. With the exception of two accessions, DaZiJie and DaZiMian, genetic similarity coefficients among all accessions ranged from 0.58 to 0.87 suggesting high level of genetic variation in the G. arboreum collections. The UPGMA dendrogram constructed from genetic similarity coefficients revealed positive correlation between cluster groupings and geographic distances. In addition, comparison of the microsatellite amplification profiles of the diploid G. arboreum and tetraploid Gossypium hirsutum L. found that size distribution of amplified products in G. arboreum was dispersive and that of G. hirsutum was relatively concentrated. The information on the genetic diversity and SSR fingerprinting from this study is useful for developing mapping populations for constructing diploid cotton genetic linkage map and tagging economically important traits.

... 以上研究结果与Liu等^[29]的观点一致, 即SSR聚类结果与地理分布一致, 遗传相似系数揭示种群之间的正相关关系 ...

2007

0.0

... 目前已在油菜^[30]、大豆^[31]构建分子身份证, 在甜瓜^[32]上建立指纹图谱QR编码, 在水稻^[33]上提出基于SSR分子指纹和商品信息构建品种身份证的新思路 ...

2009

0.0

1.667

高运来, 朱荣胜, 刘春燕, 李文福, 蒋洪蔚, 李灿东, 姚丙晨, 胡国华, 陈庆山. 黑龙江部分大豆品种分子ID的构建. 作物学报, 2009, 35: 211-218

Gao

Y L

, Zhu

R S

, Liu

C Y

, Li

W F

, Jiang

H W

, Li

C D

, Yao

B C

, Hu

G H

, Chen

Q S.

Establishment of molecular ID in soybean varieties in Heilongjiang, China. Acta Agron Sin, 2009, 35: 211-218 (in Chinese with English abstract)

A total of 1 300 cultivars have been bred in China from 1923 to 2005. To well evaluate and use the cultivars, it is necessary to identify scientifically and accurately them by DNA molecular markers due to traditional identification methods can no longer meet requirements. In the paper, 83 cultivars developed by 13 breeding units from six temperature summation zone in Heilongjiang were investigated, 157 polymorphic alleles were detected with 43 SSR primers covering 19 linkage groups of soybean genetic map, for each primer, 2 to 7 allele variations were detected in all cultivars, and 3.65 on average. According to fragment size of allele variation, the data numeralized from the PAGE bands were analyzed by the software ID Analysis 1.0. Nine primers (Satt100, Sat_218, Satt514, Satt551, Satt380, Satt193, Satt191, Satt442, and Sat_084) could be used to identify all 83 soybean cultivars. The PAGE pattern of each cultivar amplified with the 9 primers could be used as a molecular ID of the cultivars. A set of molecular ID were established for 83 soybean cultivars in Heilongjiang.

1College of agriculture, Northeast Agricultural University, Harbin 150030,China;2The Crop Research and Breeding Center of Land-Reclamation,Harbin 150090, China; 3The National Research Center of Soybean Engineering and technology, Harbin 150050,China;4College of Science, Northeast Agricural University, harbin 150030, Heilongjiang, China

以黑龙江13个育种单位6个积温带的83份大豆品种为材料, 选择分布在大豆基因组19个连锁群的43对SSR引物进行检测, 共检测出等位变异157个, 每个引物检测到的等位变异数变化范围为2~7个, 平均为3.65个。将聚丙烯酰胺凝胶电泳得到的谱带统计结果根据等位变异的片段大小数字化, 用自行编制的ID Analysis 1.0软件进行数据分析。结果表明, 仅需9对引物(Satt100、Sat_218、Satt514、Satt551、Satt380、Satt193、Satt191、Satt442、Sat_084)可将83份参试大豆品种完全区分开。构建了一套黑龙江省大豆品种的分子ID。

2012

0.0

1.889

宋海斌, 崔喜波, 马鸿艳, 朱子成, 栾非时. 基于SSR标记的甜瓜品种(系)DNA 指纹图谱库的构建. 中国农业科学, 2012, 45: 2676-2689

Song

H B

, Cui

X B

, Ma

H Y

, Zhu

Z C

, Luan

F S.

Construction of DNA fingerprint database based on SSR marker for varieties (lines) of Cucumis melo L. Sci Agric Sin, 2012, 45: 2676-2689 (in Chinese with English abstract)

【Objective】 DNA fingerprinting database of varieties of melon (Cucumis melo L.) was constructed and the identification method was obtained. 【Method】The simple sequence repeat (SSR) primers were selected using 20 representative melon varieties (lines) for establishing fingerprints of 105 different varieties (lines). 【Result】Out of the 1 219 pairs of SSR primers, 18 pairs were screened to generate polymorphic fingerprints among 105 varieties or inbred lines. The results showed that each pair of the primers could detect 4-14 polymorphic bands with an average of 9. The average polymorphism information content (PIC) was 0.68 with a range of 0.55-0.82. The genetic similarity coefficient of the 105 accessions ranged from 0.70 to 0.99. The fingerprint data of all the 18 pairs of core SSR primers were employed to establish the fingerprinting database to distinguish the 105 samples and every accession had its unique fingerprinting. 【Conclusion】 The results indicated that SSR marker is suitable to construction of DNA fingerprinting database of melon varieties and the database could provide reference for melon identification.

1.东北农业大学园艺学院，哈尔滨 150030 2.黑龙江傲龙农业有限公司，哈尔滨150020

【目的】建立甜瓜品种的DNA指纹图谱数据库，实现对甜瓜品种进行快速、准确的鉴定。【方法】应用SSR分子标记技术，首先利用20份具有代表性的甜瓜品种（系）筛选SSR引物，然后对105份不同甜瓜品种（系）进行指纹图谱的构建。【结果】从1 219对SSR引物中筛选出18对引物为105份材料形成了多态性的指纹图谱，其中每对引物可以检测到4—14条数目不等的多态性条带，平均为9条；多态性信息含量（PIC）平均为0.68，变化范围为0.55—0.82。105份材料间的相似系数为0.70—0.99。利用这18对SSR核心引物构建的指纹图谱库能够有效区分所有供试材料，并且为每份材料建立了一份独特的指纹图谱。【结论】SSR标记适于构建甜瓜品种（系）的DNA指纹图谱库，可为甜瓜品种鉴定提供依据。

2014

0.0

1.667

陆徐忠, 倪金龙, 李莉, 汪秀峰, 马卉, 张小娟, 杨剑波. 利用SSR分子指纹和商品信息构建水稻品种身份证. 作物学报, 2014, 40: 1-9

X Z

, Ni

J L

, Li

, Wang

X F

, Ma

, Zhang

X J

, Yang

J B.

Construction of rice variety based on ID used SSR fingerprint and commodity information. Acta Agron Sin, 2014, 40: 1-9 (in Chinese with English abstract)

For increasing convenience of the rice seed quality tracking and management, we selected a set of 12 rice SSR markers (one marker for each chromosome) based on previous research. The fluorescent marker analysis was performed on 127 released and popularized authorized (or main plant) rice varieties in Anhui Province. Then, the rice variety ID was constructed by the SSR fingerprint with corresponded commodity information. The rice variety ID included three components: part 1 was recognized as commodity code, which reflected crop species, cultivar, breeding (or approval) region and year; part 2 was recognized as fingerprint code, which contained the variety DNA fingerprint information; part 3 was recognized as complement code (specific gene identification code), reflecting specific genetic information of cultivars. We also described the rice variety ID in the form of bar code or QR code, which might be favorable for rice seed identification and traceability management.

1 Institute of Rice Research, Anhui Academy of Agricultural Sciences, Hefei 230031, China; 2.Institute of Cotton Research, Anhui Academy of Agricultural Sciences, Anqing 246003, China

为方便水稻种子质量的追溯和管理，提出基于SSR分子指纹和商品信息构建水稻品种身份证的新思路。在前期研究基础上，进一步确定了由12个水稻SSR标记构成的核心标记组(每条染色体1个)，对127份安徽省审定(或主推)水稻品种进行荧光标记分析，构建供试水稻品种分子指纹，在此基础上，再与品种基本商品信息相结合，并进行数字化编码，构建了127份水稻的品种身份证。该身份证号码组成的第1部分为商品码，代表品种的基本商品信息，包括品种类别、品种选育(或审定)的区域和时间等；第2部分为指纹码，代表水稻品种的分子指纹信息；第3部分为补充码(或特异基因识别码)，代表品种的特异基因信息。还对127份水稻品种身份证进行了条码表述(一维码和二维码)，以便水稻品种种子的身份标识和溯源管理。