王立新,张小军,史星雲,高华,赵政阳

• 1:西北农林科技大学园艺学院
• 2:陕西省苹果工程技术研究中心

摘要(Abstract)：

【目的】通过构建苹果栽培品种的SSR标记指纹图谱数据库,以实现对苹果品种的快速、准确鉴定。【方法】以国内外40个主要栽培品种为试验材料,选用均匀分布于17条染色体上的144对SSR引物进行PCR扩增并采用6%变性聚丙烯酰胺凝胶电泳与SSR荧光标记毛细管电泳相结合的方法进行检测。【结果】结果表明,从144对SSR引物中筛选出了35对多态性较高、谱带清晰的引物,35对引物共检测到314个等位基因,每对SSR引物扩增出等位基因数3～15个,平均8.97个;多态性信息含量指数(PIC)变化为0.455 3～0.834 6,平均为0.692 0;各个位点的杂合度在0.475 0～1.000 0,平均为0.794 3;标记系数(MI)平均为6.208 0,其中CH04c10引物对苹果基因组DNA变异检测最有效,其多态性信息含量指数和标记指数均最高,分别为0.834 6和12.519 0。以CH03d07、NZ02b1和CH05c04这3对引物进行组合鉴定,即可将40个苹果品种完全区分开。与6%变性聚丙烯酰胺凝胶电泳检测方法相比,荧光标记毛细管电泳检测能更准确读出目标片段的大小,检测效果更为理想,更适合于DNA指纹图谱数据库的建立。【结论】将2种检测方法结合构建苹果品种DNA指纹图谱,结果更准确、更可靠。

关键词(KeyWords)： 苹果;SSR标记;荧光标记;指纹图谱

Abstract:

Keywords:

基金项目(Foundation): 公益性行业(农业)科研专项(200903008-11);; 陕西省科技统筹创新工程计划项目(2011KTZB02-02-01);; 国家现代苹果产业技术体系(CARS-28)

作者(Author): 王立新,张小军,史星雲,高华,赵政阳

DOI: 10.13925/j.cnki.gsxb.2012.06.009

参考文献(References)：

• [1]WANG Zhuang-wei,ZHAO Mi-zhen,YUAN Ji,WU Wei-min,QIAN Ya-ming.Establishment of SSR fingerprinting database for 38cultivars of strawberry(Fragaria ananassa)native to Europe andAmerica[J].Journal of Fruit Science,2011,28(6):1032-1037.王壮伟,赵密珍,袁骥,吴伟民,钱亚明.38个欧美草莓栽培品种SSR指纹图谱的构建[J].果树学报,2011,28(6):1032-1037.
• [2]DONG Yu,ZHANG Wei,YUAN Jian-xia,ZHAO Ya-juan,ZHANGBo,LEI Xiao-ping,LI Chao,WANG Xiao-mei,HAN Tao.The in-ternational development trend analysis of crop molecular designbreeding.[Publication is unknown],2009,[http://ir.las.ac.cn/handle/12502/2838]董瑜,张薇,袁建霞,赵亚娟,张博,雷孝平,李超,王晓梅,韩涛.作物分子设计育种的国际发展态势分析[OL/J].2009,[http://ir.las.ac.cn/handle/12502/2838]
• [3]WANG Feng-ge,ZHAO Jiu-ran,GUO Jing-lun,LIU Long-zhou.Series of research on establishing DNA fingerprinting pool of chinesenew maize cultivars:I.The establishment of a standard SSR systemfitting for maize cultivars identification[J].Journal of Maize Science,2003,11(1):3-6.王凤格,赵久然,郭景伦,刘龙洲.中国玉米新品种DNA库建立系列研究:I.玉米品种纯度及真伪鉴定中SSR标准实验体系的建立[J].玉米科学,2003,11(1):3-6.
• [4]GAO Hua,DNA fingerprinting and genetic diversity in apple culti-vars[D].Yangling:Northwest A&F University,2010.高华.苹果栽培品种的DNA指纹图谱构建及遗传多样性分析[D].杨凌:西北农林科技大学,2010.
• [5]GAO Yuan,WANG Kun,TIAN Lu-ming,CAO Yu-fen,LIU Feng-zhi.Identification of apple cultivars by TP-M13-SSR technique[J].Journal of Fruit Science,2010,27(5):833-837.高源,王昆,田路明,曹玉芬,刘凤之.应用TP-M13-SSR技术鉴定苹果品种[J].果树学报,2010,27(5):833-837.
• [6]GAO Yuan,WANG Kun,TIAN Lu-ming,CAO Yu-fen,LIU Feng-zhi.TP-M13-SSR technique and its applications in analysis of ge-netic diversity for apple germplasm resources[J].Journal of Plant Ge-netic Resources,2011,12(2):228-233.高源,王昆,田路明,曹玉芬,刘凤之.TP-M13-SSR技术及其在苹果种质资源遗传多样性研究中的应用[J].植物遗传资源学报,2011,12(2):228-233.
• [7]HAO Chen-yang,WANG Lan-fen,JIA Ji-zeng,DONG Yu-shen,ZHANG Xue-yong.Comparison of fluorescence and silver-stainingdetection systems of microsatellite markers[J].Acta AgronomicaAinica,2005,31(2):144-149.郝晨阳,王兰芬,贾继增,董玉琛,张学勇.SSR标记和银染技术的比较分析[J].作物学报,2005,31(2):144-149.
• [8]TAKEZAKI N,NEI M.Genetic distances and reconstruction of phy-logenetic trees from microsatellite DNA[J].Genetics,1996,144:389-399.
• [9]GAO Yuan,LIU Feng-zhi,CAO Yu-fen,WANG Kun.Analysis ofgenetic relationship for Malus germplasm resources by SSR Markers[J].Journal of Fruit Science,2007,24(2):129-134.高源,刘凤之,曹玉芬,王昆.苹果属种质资源亲缘关系的SSR分析[J].果树学报,2007,24(2):129-134.
• [10]WANG Ai-de,LI Tian-zhong,XU Xue-feng,HAN Zhen-hai.SSRanalysis for apple cultivars[J].Acta Horticulturae Sinica,2005,32(5):875-877.王爱德,李天忠,许雪峰,韩振海.苹果品种的SSR分析[J].园艺学报,2005,32(5):875-877.
• [11]TAUTZ D.Hypervariability of simple sequences as a general sourcefor polymorphic DNA markers[J].Nucleic Acids Research,1989,17(16):6463-6471.
• [12]GUIFORD P,PRAKASH S,ZHU J M,RIKKERINK E,GARDINERS,BASSETT H,FORSTER R.Microsatellites in Malus×domestica(apple):abundance,polymorphism and cultivar identification[J].Theoretical and Applied Genetics,1997,94:249-254.
• [13]LUIS G,CRISTINA M,OLIVEIRA.Molecular characterization ofcultivars of apple(Malus×domestica Borkh.)using microsatellite(SSR and ISSR)markers[J].Euphytica,2001,122:81-89.
• [14]CAI Qing,JIANG Li-jie,ZHANG Xiao-ming,YAN Guo-hua,ZHANG Kai-chun,CAO Yu-fen,MA Huan-pu.Identification ofapple varieties with SSR molecular markers[J].Chinese AgriculturalScience Bulletin,2007(7):129-134.蔡青,姜立杰,张晓明,闫国华,张开春,曹玉芬,马焕普.苹果主栽品种的SSR分子标记鉴别[J].中国农学通报,2007(7):129-134.
• [15]LIEBHARD R,GIANFRANCESCHI L,KOLLER B,RYDER C D,TARCHINI R,VAN DE,WEG E,GESSLER C.Development andcharacterisation of 140 new microsatellites in apple(Malus×domes-tica Borkh.)[J].Molecular Breeding,2002,10:217-241.
• [16]CHEN Chen,ZHANG Xing-tao,CHENG Fei,ZHUANG Mu,LIUYu-mei,YANG Li-mei,ZHANG Yang-yong,FANG Zhi-yuan.Establishment of SSR fingerprinting on autumn cabbage hybrids andtheir parents[J].Acta Horticulturae Sinica,2011,38(1):159-164.陈琛,张兴桃,程斐,庄木,刘玉梅,杨丽梅,张扬勇,方智远.秋甘蓝品种的SSR指纹图谱的构建[J].园艺学报,2011,38(1):159-164.
• [17]AI Cheng-xiang,QIN Zhi-hua,TAO Ji-han,WANG Chang-jun.SSR fingerprints and genetic variations of the 32 persimmon majorcultivars[J].Acta Botanica Boreali-Occidentalia Sinica,2011,31(11):2185-2191.艾呈祥,秦志华,陶吉寒,王长君.32个柿主栽品种SSR构建及遗传变异分析[J].西北植物学报,2011,31(11):2185-2191.
• [18]CHENG Ben-yi,XIA Jun-hui,GONG Jun-yi,YANG Shi-hua.Application of capillary electrophoresis detection with fluorescentSSR markers in rice DNA fingerprint identification[J].China Jour-nal Rice Science,2011,25(6):672-676.程本义,夏俊辉,龚俊义,杨仕华.SSR荧光标记毛细管电泳检测法在水稻DNA指纹鉴定中的应用[J].中国水稻科学,2011,25(6):672-676.