轮枝镰孢SSR标记开发及在玉米分离群体遗传多样性分析中的应用

doi:10.3864/j.issn.0578-1752.2012.01.007

摘要/Abstract

摘要： 【目的】开发轮枝镰孢（Fusarium verticillioides）的SSR标记，为轮枝镰孢遗传多样性研究提供技术支持。【方法】利用Fastpcr在轮枝镰孢基因组中查找符合条件的SSR位点，采用Primer5.0软件设计引物，利用NTSYS及Popgene分析来自玉米的轮枝镰孢单孢分离物群体的PCR扩增结果。【结果】共设计有效扩增SSR引物158对，经筛选，109对（69.0%）可扩增出2条及以上的条带，其中55对引物（34.8%）多态性较好，可扩增出3条及以上的条带。从11条已组装的轮枝镰孢染色体上各选出2对多态性引物，计22对引物，对66株轮枝镰孢玉米分离物进行扩增，共获得125个等位变异，变异范围为2—11个，平均为5.68个。轮枝镰孢玉米分离物之间Nei’s基因多样性指数为0.1139—0.8687，平均为0.6199，表现出较高的遗传多样性。【结论】基于轮枝镰孢基因组序列开发的SSR标记具有很好的多态性，可用于轮枝镰孢的遗传多样性分析。用22对SSR引物扩增，以遗传相似系数0.3进行划分，可将66株轮枝镰孢玉米分离物分为3群；中国轮枝镰孢玉米分离物的遗传多样性与地理分布无相关性。

关键词: 轮枝镰孢, 简单串联重复序列, 玉米分离物, 遗传多样性

Abstract: 【Objective】 The objective of this study is to analyze genetic diversity of Fusarium verticillioides and to develop SSR markers for this fungus. 【Method】 Fastpcr was used for searching SSRs in genomic sequence of F. verticillioides. The sequences including suitable SSRs were selected to design primers using Primer5.0. The NTSYS and Popgene programs were used to analyze genetic diversity of maize isolates of F. verticillioides. 【Result】 Among the 158 primer pairs designed, 109 pairs (69.0%) were able to amplify 2 or more alleles against discretionary F. verticillioides isolates originating from single conidium. Of them 55 pairs (34.8%) amplified 3 or more alleles. In 11 chromosomes assembled 22 primer pairs, two pairs for one chromosome, were selected for genetic diversity analysis of 66 isolates of F. verticillioides from maize. In 22 SSR loci, a total of 125 alleles (2 to 11 alleles amplyfied by per primer pair) were detected with an average of 5.68 alleles. The Nei’s gene diversity, ranged from 0.1139 to 0.8687 with an average mean of 0.6199, indicated high genetic diversity in F. verticillioides isolates from various geographic regions.【Conclusion】Based on genome information of F. verticillioides, 109 SSR markers were developed and showed high polymorphism and high diversity among F. verticillioides isolates. By amplyfying with 22 SSR primer pairs, 66 isolates of F. verticillioides from maize were seperated into 3 groups on 0.3 coefficient. The relationship between genetic diversity and geographical distribution did not show correlation in isolates of F. verticillioides from maize in China.

Key words: Fusariumverticillioides, simplesequancerepeat(SSR), pathogenisolatefrommaize, geneticdiversity

任旭, 朱振东, 李洪杰, 段灿星, 王晓鸣. 轮枝镰孢SSR标记开发及在玉米分离群体遗传多样性分析中的应用[J]. 中国农业科学, 2012, 45(1): 52-66.

REN Xu, ZHU Zhen-Dong, LI Hong-Jie, DUAN Can-Xing, WANG Xiao-Ming. SSR Marker Development and Analysis of Genetic Diversity of Fusarium verticillioides Isolated from Maize in China[J]. Scientia Agricultura Sinica, 2012, 45(1): 52-66.

0
/ / 推荐

导出引用管理器 EndNote|Reference Manager|ProCite|BibTeX|RefWorks

链接本文: https://www.chinaagrisci.com/CN/10.3864/j.issn.0578-1752.2012.01.007

https://www.chinaagrisci.com/CN/Y2012/V45/I1/52

参考文献

[1]Görtz A, Oerke E C, Steiner U, Waalwijk C, de Vries I, Dehne H W. Biodiversity of Fusarium species causing ear rot of maize in Germany. Cereal Research Communications, 2008, 36(Suppl. B): 617-622.

[2]Krnjaja V, Levic J, Tomic Z, Nesic I Z, Stojanovic L, Trenkovski S. Dynamics of incidence and frequency of populations of Fusarium species on stored maize grain. Biotechnology in Animal Husbandry, 2007, 23(5/6): 589-600.

[3]Munkvold G P, Hellmich R L, Rice L G. Comparison of fumonisin concentrations in kernels of transgenic Bt maize hybrids and nontransgenic hybrids. Plant Disease, 1999, 83(2): 130-138.

[4]马秉元, 龙书生, 李多川, 李亚玲. 陕西省玉米穗粒腐病的病原菌鉴定及各分离菌分布频率. 西北农业大学学报, 1995, 23(Suppl.): 98-103.

Ma B Y, Long S S, Li D C, Li Y L. A study of the identification of corn kernel or ear rot pathogens in Shaanxi Province. Acta University of Agricultural Boreali-occidentalis, 1995, 23(Suppl.): 98-103. (in Chinese)

[5]陈晓娟, 文成敬. 四川省玉米穗腐病研究初报. 西南农业大学学报, 2002, 24(1): 21-23, 25.

Chen X J, Wen C J. Preliminary study of maize ear rot in Sichuan. Journal of Southwest Agricultural University, 2002, 24(1): 21-23, 25. (in Chinese)

[6]Murillo-Williams A, Munkvold G P. Systemic infection by Fusarium verticillioides in maize plants grown under three temperature regimes. Plant Disease, 2008, 92(12): 1695-1700.

[7]Wilke A L, Bronson C R, Tomas A, Munkvold G P. Seed transmission of Fusarium verticillioides in maize plants grown under three different temperature regimes. Plant Disease, 2007, 91(9): 1109-1115.

[8]宋立秋, 魏利民, 王振营, 何康来, 丛斌. 亚洲玉米螟与串珠镰孢菌复合侵染对玉米产量损失的影响. 植物保护学报, 2009, 36(6): 487-490.

Song L Q, Wei L M, Wang Z Y, He K L, Cong B. Effect of infestation by the Asian corn borer together with Fusarium verticillioides on corn yield loss. Acta Phytophylacica Sinica, 2009, 36(6): 487-490. (in Chinese)

[9]Gohari A M, Javan-Nikkhah M, Hedjaroude G A, Abbasi M, Rahjoo V, Sedaghat N. Genetic diversity of Fusarium verticillioides isolates from maize in Iran based on vegetative compatibility grouping. Journal of Plant Pathology, 2008, 90(1): 113-116.

[10]Chulze S N, Ramirez M L, Torres A, Leslie J F. Genetic variation in Fusarium section Liseola from no-till maize in Argentina. Applied and Environmental Microbiology, 2000, 66(12): 5312-5315.

[11]Danielsen S, Meyer U M, Jensen D F. Genetic characteristics of Fusarium verticillioides isolates from maize in Costa Rica. Plant Pathology, 1998, 47(5): 615-622.

[12]Reynoso M M, Chulze S N, Zeller K A, Torres A M, Leslie J F. Genetic structure of Fusarium verticillioides populations isolated from maize in Argentina. European Journal of Plant Pathology, 2009, 123(2): 207-215.

[13]Mohammadi A, Mofrad N N. Investigation on genetic diversity of Fusarium verticillioides isolated from corn using vegetative compatibility groups and relation of VCGs to the pathogenecity. Journal of Agricultural Technology, 2011, 7(1): 143-148.

[14]Krnjaja V S, Levi J T, Stankovi S Z, Tomic Z M. Pathogenicity and diversity of vegetative compatibility of Fusarium verticillioides. Zbornik Matice Srpske za Prirodne Nauke/Matica Srpska Proceedings for Natural Sciences, 2007, 113: 103-111.

[15]Ono E Y S, Fungaro M H P, Sofia S H, Miguel T A, Sugiura Y, Hirooka E Y. Fusarium verticillioides strains isolated from corn feed: characterization by vumonisin pr?duction and RAPD fingerprinting. Brazilian Archives of Biology and Technology, 2010, 53(4): 953-960.

[16]æ周晓云. 李瑟组镰孢霉的遗传研究[D]. 南京: 匪京农业大学, 2001.

Zhou X Y. Gene²ics of Fusarium section Liseola[D]. Nanjing: Nanjing Agricultural University, 2001. (in Chinese)

[17]da Silva V N, Fernandes F M C, Cortez A, Ribeiro D H B, de Almeida A P, Hassegawa R H, Corrêa B. Characterization and genetic variability of Fusarium verticillioides strains isolated from corn and sorghum in Brazil based on fumonisins production, microsatellites, mating type locus, and mating crosses. Canadian Journal of Microbiology, 2006, 52(8): 798-804.

[18]Moretti A, Mulè G, Susca A, González-Jaén M T, Logrieco A. Toxin profile, fertility and AFLP analysis of Fusarium verticillioides from banana fruits. European Journal of Plant Pathology, 2004, 110(5): 601-609.

[19]Rocha L O, Reis G M, da Silva V N, Braghini R, Teixeira M M G, Corrêa B. Molecular characterization and fumonisin production by Fusarium verticillioides isolated from corn grains of different geographic origins in Brazil. International Journal of Food Microbiology, 2011, 145(1): 9-21.

[20]Rahjou V, Zad J, Nikkhah M J, Bihamta M R, Okhovat S M, Mirzadi G A, Elameen A A H, Klemsdal S. S. Study of genetic variation in isolates of Fusarium verticillioides (Sacc.) Nirenberg, the causal agent of Fusarium ear rot of corn using AFLP markers. Seed and Plant, 2008, 24(3): 457-476.

[21]Scott J B, Chakraborty S. Identification of 11 polymorphic simple sequence repeat loci in the phytopathogenic fungus Fusarium pseudograminearum as a tool for genetic studies. Molecular Ecology Resources, 2008, 8(3): 628-630.

[22]Vogelgsang S, Widmer F, Jenny E, Enkerli J. Characterisation of novel Fusarium graminearum microsatellite markers in different Fusarium species from various countries. European Journal of Plant Pathology, 2009, 123(4): 477-482.

[23]Singh R, Sheoran S, Sharma P, Chatrath R. Analysis of simple sequence repeats (SSRs) dynamics in fungus Fusarium graminearum. Bioinformation, 2011, 5(10): 402-404.

[24]张昊, 张争, 许景升, 徐进, 张丽勍, 潘哲超, 田茜, 冯洁. 一种简单快速的赤霉病菌单孢分离方法——平板稀释画线分离法. 植物保护, 2008, 34(6): 134-136.

Zhang H, Zhang Z, Xu J S, Xu J, Zhang L Q, Pan Z C, Tian Q, Feng J. A rapid and simple method for obtaining single-spore isolates of Fusarium species-agar dilution lineation separation. Plant Protection, 2008, 34(6): 134-136. (in Chinese)

[25]Leslie J F, Summerell B A. The Fusarium Laboratory Manual. Iowa: Blackwell Publishing, 2006.

[26]Stewart C N Jr, Via L E. A rapid CTAB DNA isolation technique useful for RAPD fingerprinting and other PCR applications. Bio Techniques, 1993, 14(5):748-749.

[27]Mulè G, Susca A, Stea G, Moretti A. A species-specific PCR assay based on the calmodulin partial gene for identification of Fusarium verticillioides, F. proliferatum and F. subglutinans. European Journal of Plant Pathology, 2004, 110(5): 495-502.

[28]Ma L J, van der Does H C, Borkovich K A, Coleman J J, Daboussi M J, Pietro A D, Dufresne M, Freitag M, Grabherr M, Henrissat B, Houterman P M, Kang S, Shim W B, Woloshuk C, Xie X, Xu J R, Antoniw J, Baker S E, Bluhm B H, Breakspear A, Brown D W, Butchko R A E, Chapman S, Coulson R, Coutinho P M, Danchin E G J, Diener A, Gale L R, Gardiner D M, Goff S, Hammond-Kosack K E, Hilburn K, Hua-Van A, Jonkers W, KazanK, Kodira C D, Koehrsen M, Kumar L, Lee Y H, Li L, Manners J M, Miranda-Saavedra D, Mukherjee M, Park G, Park J, Park S Y, Proctor R H, Regev A, Ruiz-Roldan M C, Sain D, Sakthikumar S, Sykes S, Schwartz D C, Turgeon B G, Wapinski I, Yoder O, Young S, Zeng Q, Zhou S, Galagan J, Cuomo C A, Kistler H C, Rep M. Comparative genomics reveals mobile pathogenicity chromosomes in Fusarium. Nature, 2010, 464(7287): 367-373.

[29]Xu J R, Yan K, Dickman M B, Leslie J F. Electrophoretic karyotypes distinguish the biological species of Gibberella fujikuroi (Fusarium section Liseola). Molecular Plant Microbe Interactions, 1995, 8(1): 74-84.

[30]Kalendar R, Lee D, Schulman A H. FastPCR software for PCR primer and probe design and repeat search. Genes, Genomes and Genomics, 2009, 3(1): 1-14.

[31]徐静静, 王晓鸣, 段灿星, 武小菲, 朱振东. 大豆疫霉基因组SSR标记开发. 中国农业科学, 2009, 42(9): 3112-3122.

Xu J J, Wang X M, Duan C X, Wu X F, Zhu Z D. Development of genomic SSR markers for Phytophthora sojae. Scientia Agricultura Sinica, 2009, 42(9): 3112-3122. (in Chinese)

[32]Dyer A T, Leonard K J. Contamination, error, and nonspecific molecular tools. Phytopathology, 2000, 90(6): 565-567.

[33]Farrokhi-Nejad R, Leslie J F. Vegetative compatibility group diversity within populations of Fusarium moniliforme isolated from corn seed. Phytopathology, 1990, 80: 1043.

[34]Wolf P G, Soltis P S. Estimates of gene flow among populations, geographic races, and species in the Ipomopsis aggregata complex. Genetics, 1992, 130(3): 639-647.

[35]Souza F L, Cunha A F, Oliveira M A, Pereira G A G, dos Reis S F. Estimating dispersal and gene flow in the neotropical freshwater turtle Hydromedusa maximiliani (Chelidae) by combining ecological and genetic methods. Genetics and Molecular Biology, 2002, 25(2): 151-155.