西瓜枯萎病菌和品种的遗传多样性研究
摘要
西瓜是世界上的重要水果之一，尤其在夏季水果中占有十分突出的地位，其栽培面积与总产量均在世界十大水果之列。而西瓜枯萎病的发生严重威胁着西瓜的产量和品质，是西瓜生产上的主要病害。理论研究和生产实践证明，培育西瓜抗病品种是解决西瓜枯萎病病害最经济有效途径。而抗病品种的培育涉及到病菌分化、抗源的鉴定和收集、品种的抗性遗传和抗病机制等的研究。因此，开展西瓜枯萎病菌和品种的遗传多样性研究，能够为西瓜抗病育种和品种的科学利用提供可靠的理论基础和方法。本研究旨在研究西瓜枯萎病菌和西瓜品种的遗传分化，并寻找病菌遗传分化和致病性分化、西瓜品种遗传多样性和抗病性之间的关系。
本研究是在枯萎病菌专化型、致病型鉴定和西瓜品种抗病性鉴定的基础上，一方面以河北省西瓜主产区的47个枯萎病菌和来自北京的1号生理小种、新疆的0号、2号生理小种为试材，采用RAPD分子标记技术，对西瓜枯萎病菌的遗传分化进行了分析和讨论；另一方面以48个来源和抗性不同的西瓜品种为试材，采用AFLP分子标记技术，对西瓜品种的遗传多样性进行了研究。
1．采用完全随机试验，以病菌FON6为试材，建立了适合西瓜枯萎病菌的RAPD反应体系：随机引物浓度为10μmol／L，模板DNA浓度为40ng，Taq DNA聚合酶1.25U，10×buffer 2.5μL，2mmol／L dNTPs 1μL；同时对RAPD扩增反应中适宜的退火温度进行了探索，最终确定为37℃。
2．对50个西瓜枯萎病菌进行遗传多样性分析。结果表明，50个供试菌系两两之间的相似系数变化在0.45～0.93之间，平均遗传相似系数为0.71，可见西瓜枯萎病菌群体内存在较丰富的遗传变异。基于RAPD聚类分析表明，50个菌系可划分为6个RAPD群(RAPD groups,简称ROs)，即RGⅠ、RGⅡ、RGⅢ、RGⅣ、RGⅤ和RGⅥ。其中RGⅥ又可划分为7个亚群(RAPD subgroups，简称RSGs)，分别为RSG1、RSG2、RSG3、RSG4、RSG5、RSG6和RSG7。不同的RGs或RSGs含有的菌系数目不同，RGⅠ和RGⅡ仅含有一个菌系，RGⅢ和RGⅣ分别包括两个菌系，RGⅣ包括5个菌系，其余的菌系属于RGⅥ；RSG1至RSG5各包括一个菌系，RSG4、RSG5各包括两个菌系；RSG5包括19个菌系，且这19个菌系之间的亲缘关系变异较大；RSG6共包括13个菌系，各菌系间的遗传变异程度亦有所不同。本研究结果表明：(1)RAPD类群(或亚群)与病菌致病型间没有明显的相关性；(2)RAPD类群(或亚群)的划分与菌系地理来源亦无明显的相关性。
3．11对引物对河北省48个西瓜品种进行的AFLP分析表明，48个品种之间的相似系数变化在0.32～0.95之间，平均相似系数为0.82，相似系数变化幅度大是品种特大京欣作用的结果，该品种的遗传背景与其他47个品种明显不同，若剔除特大京欣，则其余47个品种间的相似系数变化范围为0.59～0.95，表明
西瓜品种之间遗传基础狭窄。聚类分析结果表明，48个品种被划分为6个类群
(AGs)，即类群AGI、AG 11、AGm、AGW、AGV和AGVI。其中AGI、
AGn和AGnl各包括1个品种，AGIV包括3个品种，AGV包括20个品种，
AGVI包括22个品种。类群AGV又可划分为3个亚群(A SGs)，即ASGI、
ASGll和ASGm。基于AFLP多态性带划分的品种类群与品种的抗病性鉴定
结果不能很好的统一起来。
本研究的结果建立了西瓜枯萎病菌RAPD体系;明确了西瓜枯萎病菌和西
瓜品种的遗传多样性;探讨了西瓜枯萎病菌RAPD类群与不同生理小种之间
的关系，以及西瓜品种AFLP类群与品种抗病性之间的关系。
Watermelon is one of the important fruits all over the world,and posessess a projecting position in the fruits of summer season.Its cultivating areas and total output is among the front ten fruits in the worldwide.But fusarium wilt is a main disease in the production of watermelon, which threats watermelon's production and quality seriously. Theory research and practice demonstrate that breeding the watermelon fusarium disease-resistant cultivars is a main way to resolve this problem, which includes differentiation of the isolates, identifying and collecting of resistant material,and resisrance heredity and disease-resistant mechanism. So making experiments on the genetic differentiation of Fusarium Oxysporum f. sp. niveum on the DNA level, and the genetic diversity of watermelon cultivars can provide theoretic foundation and methods to breed resistant cultivars and make full use of watermelon cultivars.In this research,we aim to study differentiation of Fusarium Oxysporum f.sp.niveum and genetic diversity o
f watermelon cultivars. Meanwhile, analyzing the relationship between differentiation and pathogenic type, and the relationship between genetic diversity and cultivars' resistant power.
Based on the identification of specialization types and pathogenic types with Fusarium Oxporum f.sp. niveum, and disease-resistant results of cultivars. On one side, with 47 isolates from the main watermelon product area of Hebei province and physiologic race 1 from Beijing and physiologic race 0 and race 2 from Xinjiang,using RAPD method, we analyzed the differentiation of Fusarium Oxysporum f.sp.niveum. On the other side, we study the genetic diversity of 48 watermelon cultivars from different orign and with different resistance using AFLP.
1.By random experiment thoroughly, with isolate FON6, we established RAPD technique system fit for Fusarium Oxporum f.sp. niveum, the concentration of random premier was 10u.mol/L, template 1.6mg/L, Taq DNA polymerase 1.25U, 10xbuffer 2.5uL, dNTPs 2mmol/uL. Meanwhile, we groped on the fitful annealing tempreture. It's 37C.
2.50 isolates were analyzed on genetic diversity. The result indicated that the
range of the similarity coefficient among the 50 isolates is from 0.45 to 0.93. And the average is 0.71. It indicated that there existed seriously genetic differentiation between these isolates of Hebei province apparently. Based on the result of RAPD clustering, 50 isolates were divided into 6 RAPD groups(RGs for short). They are RGI ,RG II,RG III,RG IV RG V and RGVI. And R.GVI was subdivided into 7 RAPD subgroups (RSGs for short). They are RSG1, RSG2, RSG3, RSG4, RSG5, RSG6, and RSG7. Different RGs or RSGs had different isolate number. RG I and RGII include only one isolate separately, RGlIIand RGV include two, RGIV include 5, others belong to RGVI.Among RSGs, from RSG1 to RSG3 include only one isolate separately, RSG4 and RSG5 include two, RSG6 include 19, and there were great differentiation among the 19 isolates, RSG7 include 13. There were different level of genetic differentiation. This study demostrates: (l)There were no correlation between RAPD groups or RAPD subgroups and watermelon fusarium wilt p
athogenic types. (2) There were also no correlation between RAPD groups or RAPD subgroups and the isolates orign.
3.With 11 paired premiers,48 watermelon cultivars were analyzed by clustering using AFLP. The result indicate that the range of the similarity coefficient among the 48 cultivars is from 0.32 to 0.95,and the average 0.82. The wide varient is only because of watermelon cultivar TeDaJingXin, which is different from other cultivars on heredity background. If ticking out TeDaJingXin, the similarity coefficient range from 0.59-0.95 among the other 47 cultivars. Which indicated that the genetic base of watermelon is narrow. By clustering, 48 cultivars were divided into 6 groups. They are group AG I AGII AGIII AG IV AG V and AGVI. AG I AG II and AGIII include only one cultivar separately, AGiV. include3, AGVinclude 20, AGVI include 22. AGVcan be subdivided into 3 subgroups. The
本研究是在枯萎病菌专化型、致病型鉴定和西瓜品种抗病性鉴定的基础上，一方面以河北省西瓜主产区的47个枯萎病菌和来自北京的1号生理小种、新疆的0号、2号生理小种为试材，采用RAPD分子标记技术，对西瓜枯萎病菌的遗传分化进行了分析和讨论；另一方面以48个来源和抗性不同的西瓜品种为试材，采用AFLP分子标记技术，对西瓜品种的遗传多样性进行了研究。
1．采用完全随机试验，以病菌FON6为试材，建立了适合西瓜枯萎病菌的RAPD反应体系：随机引物浓度为10μmol／L，模板DNA浓度为40ng，Taq DNA聚合酶1.25U，10×buffer 2.5μL，2mmol／L dNTPs 1μL；同时对RAPD扩增反应中适宜的退火温度进行了探索，最终确定为37℃。
2．对50个西瓜枯萎病菌进行遗传多样性分析。结果表明，50个供试菌系两两之间的相似系数变化在0.45～0.93之间，平均遗传相似系数为0.71，可见西瓜枯萎病菌群体内存在较丰富的遗传变异。基于RAPD聚类分析表明，50个菌系可划分为6个RAPD群(RAPD groups,简称ROs)，即RGⅠ、RGⅡ、RGⅢ、RGⅣ、RGⅤ和RGⅥ。其中RGⅥ又可划分为7个亚群(RAPD subgroups，简称RSGs)，分别为RSG1、RSG2、RSG3、RSG4、RSG5、RSG6和RSG7。不同的RGs或RSGs含有的菌系数目不同，RGⅠ和RGⅡ仅含有一个菌系，RGⅢ和RGⅣ分别包括两个菌系，RGⅣ包括5个菌系，其余的菌系属于RGⅥ；RSG1至RSG5各包括一个菌系，RSG4、RSG5各包括两个菌系；RSG5包括19个菌系，且这19个菌系之间的亲缘关系变异较大；RSG6共包括13个菌系，各菌系间的遗传变异程度亦有所不同。本研究结果表明：(1)RAPD类群(或亚群)与病菌致病型间没有明显的相关性；(2)RAPD类群(或亚群)的划分与菌系地理来源亦无明显的相关性。
3．11对引物对河北省48个西瓜品种进行的AFLP分析表明，48个品种之间的相似系数变化在0.32～0.95之间，平均相似系数为0.82，相似系数变化幅度大是品种特大京欣作用的结果，该品种的遗传背景与其他47个品种明显不同，若剔除特大京欣，则其余47个品种间的相似系数变化范围为0.59～0.95，表明
西瓜品种之间遗传基础狭窄。聚类分析结果表明，48个品种被划分为6个类群
(AGs)，即类群AGI、AG 11、AGm、AGW、AGV和AGVI。其中AGI、
AGn和AGnl各包括1个品种，AGIV包括3个品种，AGV包括20个品种，
AGVI包括22个品种。类群AGV又可划分为3个亚群(A SGs)，即ASGI、
ASGll和ASGm。基于AFLP多态性带划分的品种类群与品种的抗病性鉴定
结果不能很好的统一起来。
本研究的结果建立了西瓜枯萎病菌RAPD体系;明确了西瓜枯萎病菌和西
瓜品种的遗传多样性;探讨了西瓜枯萎病菌RAPD类群与不同生理小种之间
的关系，以及西瓜品种AFLP类群与品种抗病性之间的关系。
Watermelon is one of the important fruits all over the world,and posessess a projecting position in the fruits of summer season.Its cultivating areas and total output is among the front ten fruits in the worldwide.But fusarium wilt is a main disease in the production of watermelon, which threats watermelon's production and quality seriously. Theory research and practice demonstrate that breeding the watermelon fusarium disease-resistant cultivars is a main way to resolve this problem, which includes differentiation of the isolates, identifying and collecting of resistant material,and resisrance heredity and disease-resistant mechanism. So making experiments on the genetic differentiation of Fusarium Oxysporum f. sp. niveum on the DNA level, and the genetic diversity of watermelon cultivars can provide theoretic foundation and methods to breed resistant cultivars and make full use of watermelon cultivars.In this research,we aim to study differentiation of Fusarium Oxysporum f.sp.niveum and genetic diversity o
f watermelon cultivars. Meanwhile, analyzing the relationship between differentiation and pathogenic type, and the relationship between genetic diversity and cultivars' resistant power.
Based on the identification of specialization types and pathogenic types with Fusarium Oxporum f.sp. niveum, and disease-resistant results of cultivars. On one side, with 47 isolates from the main watermelon product area of Hebei province and physiologic race 1 from Beijing and physiologic race 0 and race 2 from Xinjiang,using RAPD method, we analyzed the differentiation of Fusarium Oxysporum f.sp.niveum. On the other side, we study the genetic diversity of 48 watermelon cultivars from different orign and with different resistance using AFLP.
1.By random experiment thoroughly, with isolate FON6, we established RAPD technique system fit for Fusarium Oxporum f.sp. niveum, the concentration of random premier was 10u.mol/L, template 1.6mg/L, Taq DNA polymerase 1.25U, 10xbuffer 2.5uL, dNTPs 2mmol/uL. Meanwhile, we groped on the fitful annealing tempreture. It's 37C.
2.50 isolates were analyzed on genetic diversity. The result indicated that the
range of the similarity coefficient among the 50 isolates is from 0.45 to 0.93. And the average is 0.71. It indicated that there existed seriously genetic differentiation between these isolates of Hebei province apparently. Based on the result of RAPD clustering, 50 isolates were divided into 6 RAPD groups(RGs for short). They are RGI ,RG II,RG III,RG IV RG V and RGVI. And R.GVI was subdivided into 7 RAPD subgroups (RSGs for short). They are RSG1, RSG2, RSG3, RSG4, RSG5, RSG6, and RSG7. Different RGs or RSGs had different isolate number. RG I and RGII include only one isolate separately, RGlIIand RGV include two, RGIV include 5, others belong to RGVI.Among RSGs, from RSG1 to RSG3 include only one isolate separately, RSG4 and RSG5 include two, RSG6 include 19, and there were great differentiation among the 19 isolates, RSG7 include 13. There were different level of genetic differentiation. This study demostrates: (l)There were no correlation between RAPD groups or RAPD subgroups and watermelon fusarium wilt p
athogenic types. (2) There were also no correlation between RAPD groups or RAPD subgroups and the isolates orign.
3.With 11 paired premiers,48 watermelon cultivars were analyzed by clustering using AFLP. The result indicate that the range of the similarity coefficient among the 48 cultivars is from 0.32 to 0.95,and the average 0.82. The wide varient is only because of watermelon cultivar TeDaJingXin, which is different from other cultivars on heredity background. If ticking out TeDaJingXin, the similarity coefficient range from 0.59-0.95 among the other 47 cultivars. Which indicated that the genetic base of watermelon is narrow. By clustering, 48 cultivars were divided into 6 groups. They are group AG I AGII AGIII AG IV AG V and AGVI. AG I AG II and AGIII include only one cultivar separately, AGiV. include3, AGVinclude 20, AGVI include 22. AGVcan be subdivided into 3 subgroups. The
引文
[1] 李天艳．国内外西瓜品种选育概况及趋势[J]．中国西瓜甜瓜，1994，(1)：10～12
[2] 中国西瓜甜瓜[M]．中国农业出版社，2000
[3] 孔庆国，于喜艳．浅谈西瓜育种成就和展望[J]．长江蔬菜，2001，(8)：26～27
[4] 郑高飞，赵天才．利用超丰F1砧木品种嫁接西瓜栽培技术[J]．农业科技通讯，2000，(6)：14～15
[5] 王鸣，张显．西瓜种植资源苗期对枯萎病抗性人工接种鉴定的研究[J]．中国西瓜甜瓜，1988，(1)：6～10
[6] 王凤升．西瓜枯萎病的发生和防治途径[J]．中国西瓜甜瓜，1988，(1)：47～48
[7] Booth C. The genus fusarium. Commonwealth Mycological institute. Kew, Sur, England, 1921
[8] Sleeth B. Fusarium niveum the cause of watermelon wilt[J]. West Va.Agric.Exp.Sta.Bull, 1935, (2): 57～23
[9] Rerd J. Studies on the fusaria which cause wilt in melons[J]. Czn, J.BOT, 1958, 36: 396～410
[10] 张兴平，王鸣．我国西瓜枯萎病生理小种分化研究初报[J]．中国西瓜甜瓜 1991，(1)：39～43
[11] Crall J M. Physiologic races in Fusarium oxysporumf.sp.niveum[J]. Phyto pathology, 1963, 853～873
[12] Cirulli M. Variation of pathogenicity in Fusarium oxysporum f.sp.niveum and resistance in watermelon caltivars[J].
[13] Netzer D. Physiological races and soil population level of Fusarium wilt of watermelon. Phytoparasitica[J]. 1976, (4): 131～136
[14] Martyn R D. Fusarium oxysporm f.sp.niveum Race2:A highly aggressive new race to the United States[J].Plant Disease, 1987, (3): 71～73
[15] 王浩波，王鸣．西瓜枯萎病菌生理小种分化研究[J]．西北农业学报，1993，2(4)：67～70
[16] 顾卫红，王燕华，宋荣浩，等．上海地区西瓜枯萎病病原菌生理小种初探[J]．上海农业学报，1994，10(3)：63～67
[17] 周凤珍，康国斌．北京地区西瓜枯萎病生理小种分化研究[J]．植物保护，1996，22(4)：14～16
[18] 王冬梅，王纯利，曲丽红，等．新疆西瓜枯萎病生理小种初探[J]．新疆农业大学学报，1998，21(2)：115～118．
[19] 马立新，高复兴，刘爱新，等．山东西瓜枯萎病菌致病性分化及品种抗性的初步研究[J]．植物保护，1993，19(4)：5～8．
[20] 郑服从，贺春萍，邱小强．海南岛西瓜、青瓜和毛瓜枯萎病菌致病性的初步研究[J]．
热带作物学报，1999，20(1)：49～53
[21] Woo S L, Zoina A, Del-Sorbo G, et al. Characterization of fusarium oxysprum f. sp. phaseoli by pathogenic races, VCGs, and RAPD[J]. Phytopathology, 1996, 86(9): 966～973
[22] Fernandez D, Assigbetse K, Dubois M P, et al. Molecular characterization of races and vegetative compatibility groups in fusarium oxysprum f. sp. vasinfectura[J]. Applied and Environmental Microbiology, 1994, 60(11): 4039～4046
[23] Wyss P, Bontante P. Amplification of genomic DNA of arbuscular mycrrhizal(AM)fungi by PCR using short arbitrary primers[J]. Mycol Res, 1993, (97): 1351～1357
[24] Tommerup I C, Barton J E, O'brien P A, Reliability of RAPD fingerprinting of three basidiomycete fungi, Laccaria[J]. Hydnangium and Rhizoctonia[J]. Mycol Res, 1995, 99(2): 179～186
[25] Leal S C M, Bertioli D J, Butt T M, et al. Characterization of isolated of the entomopathogenic fungus Metarhizium anisopliae by RAPD-PCR[J]. Mycol Res, 1994, (98): 1077～1081
[26] Valalounakis D J, Fragkiadakis G A.Genetic Diversity of Fusarium oxysporum Isolates from Cucumber: Differentiation by Pathogenicity, Vegetative Compatibility and RAPD Fingerprinting[J]. Ecology and Population Biology, 1999, 89(2): 161～168
[27] Welsh J, McClelland M. Fingerprinting genomes using PCR with arbitrary primers[J]. Nucleic Acids Research, 1990, 18:7213～7218
[28] Crowhust R N, Hawthorne B T, Rikkerink E H A, et al.Differentiation of Fusarium solani f.sp. cucurbitae race 1 and 2 by random amplification of polymorphic DNA[J].Curr.Genet.1991, (20): 391～396
[29] Assigbestse K B, Fernandez D, Doubios M P, et al. Differentiation of Fusarium oxysporium fsp. vasinfectum races on cotton by random amplified polymorphic DNA (RAPD) analysis[J]. Phytopathology, 1994, (84): 622～626
[30] 陈万权．冯洁，秦庆明．DNA分子标记在植物真菌病害研究中的应用．植物保护学报，1999，26(23)：277～282
[31] Marlatt M L, Correll J C, Kaufmann P, et al. Two genetically distance populations of fusarium oxysprum f. sp.lycopersici race3 in United states[J].Plant disease, 1996, 80(12): 1336～1342
[32] Nicholson P，Kezanoor H Z，苏海．用RAPD分析和DNA指纹鉴别玉米小斑病菌O、C、T三个小种[J]．植物病理学报，(23)：114
[33] Treena Burgess, Michael J, Brenda W. Simple Sequence Repeat Markers Distinguish among Morphotypes of Sphaeropsis sapinea. Applied and Environmental Microbiology, 2001, 67(1): 354～362
[34] Barve M P, Haware M P, Sainani M N, et al. Potential of microsatellites to distinguish four races of Fusarium oxysporum fisp.ciceri prevalent in India. Theor Appl Genet,
2001, 102: 138～147
[35] Elias K S, Zamir D, Lichtman-Pleban T, et al. Population structure of fusarium oxysporum f.sp.l ycopersicil: RFLP provide genetic evidence that vegetative compatibility groups is an indicator of evolutionary origin[J]. MPMI, 1993, 6(5): 565～572
[36] Manicom B Q, Baayen R P. Restraction fragment length polymorphisms in fusarium oxysporum f.sp. dianthi and other fusaria from dianthus species[J],Plant Pathology, 1993, 42:851～857
[37] Okoli C A N, Carder J H, Barbara D J. Restriction fragment length polymorphisms(RFLPs)and there relationships of some host-adapted isolates of Verticillium dahliae[J]. Plant Pathology, 1994, 43: 33～40
[38] Cander J H, Barbara D J. Molecular variation and restriction fragment length polymorphic (RFLPs) within and between six species of Verticillium[J]. Mycological research, 1991, 95: 935～942
[39] Boman E S, Khush G S, Nelson R J. Genetic differentiation among isolates of pyricularia grisea infecting rice and weed hosts [J]. Photopathology, 1993, 83: 393～399
[40] Milgroom M G, Lipari S E, Enno R A, et al. Comparison of genetic diversity in the chestnut blitht fungus, Cryphonectria parasitiea, from China and the U.S [J].Mycological research, 1992, 96: 114～120
[41] Vainio E J, Korhonen K, Hantula J. Genetic variation in Phlebtopsis glgantecn as detected with random amplified mierosatellite (RAMS) markers[J], Mycol Res, 1998, 102(2): 187～192.
[42] Paul W Tooley, Nichole R O'Neill, Erin D Goley, et al. assessment of diversity in Claviceps africana and other Claviceps species by RAM and AFLP analyses[J]. Phytopathology, 2000, 90(10): 1126～1130.
[43] Hantula J, Müller M M. Variation within Gremmeniella abietina in Finland and other countries as determined by Random Amplified Microsatellites (RAMS)[J]. Mycol. Res.1997, 101(2): 169～175
[44] 陈洪，朱立煌，李冬梅，等．致病性念珠菌DNA的AFLP指纹图谱[J]．科学通报，1996，41(10)：935～938
[45] Majer D, Mithen R, Lewis B G, et al. The use of AFLP fingerprinting for the detection of genetic variation in fungi[J]. Mycol Res, 1996, 100(9): 1107～1111
[46] Baayen R P, O' Donnell K, Bonants P J M, et al. Gene Geneal: ogies and AFLP Analyses in the Fusarium oxysporum Complex Identify monophyletic and nonmonophyletic formae speciales causing wilt and rot disease[J]. Phytopathology, 2000, 90(8): 891～900
[47] Folkertsma R T, Vanzandvoot P M, De Groot K E, et al. Gene pool similarities of potato cyst nematode populations assessed by AFLP analysis[J]. Mol Plant-Microbe Interact, 1996, (9): 47～54
[48] Kaplsn M, Cas well-Chen E P, Lliamson V M. Assessment of host-induced selection on three geographic isolates of Heterodera schachtii using RAPD and AFLP marker[J]. Phtopathology, 1999, 89:68～73
[49] Agus P. Genetic diversity of isolates of the leptosphaeria raaeulans species complex from Australia, Europe and North America using amplified fragment length polymorphism analysis[J].Mycological Research, 2000, 104(7): 772～781
[50] 邹亚飞．棉花黄萎病菌(Verticillium dahliae)致病型AFLP分析与分子鉴定研究[D]．重庆：西南农业大学，2001
[51] 高俊莲，陈文新，Terefe W Z，等．应用AFLP技术对斜茎黄氏根瘤菌遗传多样性的分析研究[J]．应用与生物学报，1999，5(4)：387～395
[52] 何秋月，Hei Leung，Zeigler Robert S，等．稻瘟病菌变异株的AFLP分析[J]．菌物系统，2002，21(3)：363～369
[53] Namikli F, Shiomi T, Nishi K, et al. Pathogenic and genetic in the Japanese strains of fusarium oxysporum f.sp melons[J], phatopathology, 1998, 88(8): 804～810
[54] Welsh J. Fingerprinting genomes using PCR with arbitrary primers[J]. Nucleic Acids Research, 1990, (18): 7213～7218
[55] Williams G K. DNA polymorphisms amplified by arbitrary primers are useful as genetic markers[J]. Nucleic Acids Researvh, 1990, (18): 6531～6535
[56] 马峙英，王省芬，张桂寅，等．现代生物技术在棉花黄萎病菌分化研究中的应用[J]．河北农业大学学报，1998，21(2)：102～111
[57] Crowhust R N. Differentiation of Fusarium solani fisp. cucurbitae race 1 and 2 by random amplification of polymorphic DNA[J].Curr.Genet, 1991, 20:391～396
[58] 陈万权．DNA分子标记在植物真菌病害研究中的应用[J]．植物保护学报，1999，26(3)：277～282
[59] Femandez D, Tantaoui A. Random amplified polymorphic DNA(RAPD) analysis: A tool for rapid characterization of Fusarium oxysporum f.sp.albedinis isolates[J]. PhytopatholMediterr, 1994, 33: 223～229
[60] Bentley S, Pegg K G, Dale J L. Genetic.variation among a world-wide collection of isolates of Fusarium oxysporum f.sp cubense analysed by RAPD-PCR fingerprinting[J]. Myeol.Res, 1995, 99:1378～1384
[61] Manulis S, Kogan N, Reuven M, et al. Use of the RAPD technique for identification of Fusarium oxysporumf.sp dianthi from carnation[J]. Phytopatholigy, 1994, 84:98～101
[62] Mes J J, Van Doom J, Roebroeek E J A, et al. Detection and identificatuib of Fusarium oxysporum fsp. gladioli by RFLP and RAPD analysis[J]. Modern Assays for Plant Pathogenic Fungi, 1995, 63～68
[63] Grajal-Martin M J, Simon C J, Muehlbauer F J, Use of random amplified polymorphic DNA (RAPD) to characterize race 2 of Fusarium oxysporum fsp. pisi[J]. Phytopatholigy, 1993, 83:612～614
[64] Zabeau M, Vos P. Selective restriction fragment amplification: a general method for DNA fingerprinting[J]. European Patent Application. 1993,
[65] 李珊，赵桂仿．AFLP分子标记及其应用[J]．西北植物学报，2003，23(5)：830～836
[66] 王斌，翁曼丽．AFLP的原理及其应用[J]．杂交水稻，1996，(5)：27～30
[67] 杜金友，黎裕，王天宇，等．SSR和AFLP分析玉米遗传多样性的研究[J]．华北农学报，2003，18(1)：59～63
[68] 易干军，谭卫萍，霍合强，等．龙眼品种(系)遗传多样性及亲缘关系的AFLP分析[J]．园艺学报，2003，30(3)：272～276
[69] 宗绪晓．D Vaughan．A Ka发a．等．AFLP分析小豆(Vigna angularis)种内遗传多样性[J]．作物学报，2003，29(4)：562～568
[70] 朱申龙，Morti M L，Rao R．应用AFLP方法研究中国大豆的遗传多样性[J]．浙江农业学报，1998，10(6)：302～309
[71] 贾建航，李传友，金德敏，等．AFLP技术在玉米自交系类群分析研究中的应用[J]．高技术通讯，1999，(4)：43～47
[72] 高世斌，李晚忱，荣廷昭，等．玉米骨干自交系AFLP指纹图谱鉴定[J]．四川农业大学学报，2001，19(2)：126～128
[73] 罗安定，陈守才，吴坤鑫，等．AFLP在橡胶树优异种质研究中的应用[J]．植物学报，2001，43(9)：941～947
[74] 马渐新，周荣华，董玉琛，等．小麦抗条锈病基因定位及分子标记研究进展[J]．生物技术通报，1999，(1)：1～6
[75] 黄祖六，谭学林，Tragoonrung．S．等．稻米直链淀粉含量基因座位的分子标记定位[J]．作物学报，2000，26(6)：777～782
[76] 房经贵，刘大钧，马正强．利用双杂合位点标记资料构建芒果遗传图谱[J]．分子植物育种，2003，1(3)：313～319
[77] 张蕴哲，刘红霞，邬荣领，等．毛新杨×毛白杨AFLP分子遗传图谱[J]．林业科学研究，2003，16(5)：595-603
[78] 康国斌，林德佩，王叶筠，等．西瓜枯萎病菌镰刀菌酸对西瓜苗作用机制的初步探讨[J]．植物病理学报，2000，30(4)：373～374
[79] 王浩波，王鸣，马德伟．用生物间遗传学原理研究西瓜与枯萎病菌的相互关系[J]．河北农业大学学报，1994，17(3)：29～33
[80] Kim W K. Isolation of high molecular weight DNA and double -stranded RNAs from fungi[J].Canadian Journal of Botany,1990, (68): 1898～1902
[81] Rogers S O.Extraction of DNA from Basidiomycetes for ribosomal DNA hybridizations[J].Canadian Journal of Botany, 1989, (67): 1235～1243
[82] 吴少慧，张成刚，张忠泽．RAPD技术在微生物生物多样鉴定中的应用[J]．微生物学杂志，2000，20(2)：44～47
[83] Paterson A H, Brnbaker C L, Wendel J F. A rapid method for extraction of cotton(Gossypium spp.) genomic DNA suitable for RFLP or PCR analysis[J]. Plant
Molecular Biology Reporter, 1993, 11(2): 122～127
[84] Van Toai, Martin S. Optimization of silver-staining AFLP technique for soybean[J]. SoybeanGenet, Newsl, 1996, (23): 206～209
[85] 田清震，盖钧镒，喻德跃，等．大豆DNA扩增片段长度多态性(AFLP)研究[J]．大豆科学，2000，19(3)：210～217
[2] 中国西瓜甜瓜[M]．中国农业出版社，2000
[3] 孔庆国，于喜艳．浅谈西瓜育种成就和展望[J]．长江蔬菜，2001，(8)：26～27
[4] 郑高飞，赵天才．利用超丰F1砧木品种嫁接西瓜栽培技术[J]．农业科技通讯，2000，(6)：14～15
[5] 王鸣，张显．西瓜种植资源苗期对枯萎病抗性人工接种鉴定的研究[J]．中国西瓜甜瓜，1988，(1)：6～10
[6] 王凤升．西瓜枯萎病的发生和防治途径[J]．中国西瓜甜瓜，1988，(1)：47～48
[7] Booth C. The genus fusarium. Commonwealth Mycological institute. Kew, Sur, England, 1921
[8] Sleeth B. Fusarium niveum the cause of watermelon wilt[J]. West Va.Agric.Exp.Sta.Bull, 1935, (2): 57～23
[9] Rerd J. Studies on the fusaria which cause wilt in melons[J]. Czn, J.BOT, 1958, 36: 396～410
[10] 张兴平，王鸣．我国西瓜枯萎病生理小种分化研究初报[J]．中国西瓜甜瓜 1991，(1)：39～43
[11] Crall J M. Physiologic races in Fusarium oxysporumf.sp.niveum[J]. Phyto pathology, 1963, 853～873
[12] Cirulli M. Variation of pathogenicity in Fusarium oxysporum f.sp.niveum and resistance in watermelon caltivars[J].
[13] Netzer D. Physiological races and soil population level of Fusarium wilt of watermelon. Phytoparasitica[J]. 1976, (4): 131～136
[14] Martyn R D. Fusarium oxysporm f.sp.niveum Race2:A highly aggressive new race to the United States[J].Plant Disease, 1987, (3): 71～73
[15] 王浩波，王鸣．西瓜枯萎病菌生理小种分化研究[J]．西北农业学报，1993，2(4)：67～70
[16] 顾卫红，王燕华，宋荣浩，等．上海地区西瓜枯萎病病原菌生理小种初探[J]．上海农业学报，1994，10(3)：63～67
[17] 周凤珍，康国斌．北京地区西瓜枯萎病生理小种分化研究[J]．植物保护，1996，22(4)：14～16
[18] 王冬梅，王纯利，曲丽红，等．新疆西瓜枯萎病生理小种初探[J]．新疆农业大学学报，1998，21(2)：115～118．
[19] 马立新，高复兴，刘爱新，等．山东西瓜枯萎病菌致病性分化及品种抗性的初步研究[J]．植物保护，1993，19(4)：5～8．
[20] 郑服从，贺春萍，邱小强．海南岛西瓜、青瓜和毛瓜枯萎病菌致病性的初步研究[J]．
热带作物学报，1999，20(1)：49～53
[21] Woo S L, Zoina A, Del-Sorbo G, et al. Characterization of fusarium oxysprum f. sp. phaseoli by pathogenic races, VCGs, and RAPD[J]. Phytopathology, 1996, 86(9): 966～973
[22] Fernandez D, Assigbetse K, Dubois M P, et al. Molecular characterization of races and vegetative compatibility groups in fusarium oxysprum f. sp. vasinfectura[J]. Applied and Environmental Microbiology, 1994, 60(11): 4039～4046
[23] Wyss P, Bontante P. Amplification of genomic DNA of arbuscular mycrrhizal(AM)fungi by PCR using short arbitrary primers[J]. Mycol Res, 1993, (97): 1351～1357
[24] Tommerup I C, Barton J E, O'brien P A, Reliability of RAPD fingerprinting of three basidiomycete fungi, Laccaria[J]. Hydnangium and Rhizoctonia[J]. Mycol Res, 1995, 99(2): 179～186
[25] Leal S C M, Bertioli D J, Butt T M, et al. Characterization of isolated of the entomopathogenic fungus Metarhizium anisopliae by RAPD-PCR[J]. Mycol Res, 1994, (98): 1077～1081
[26] Valalounakis D J, Fragkiadakis G A.Genetic Diversity of Fusarium oxysporum Isolates from Cucumber: Differentiation by Pathogenicity, Vegetative Compatibility and RAPD Fingerprinting[J]. Ecology and Population Biology, 1999, 89(2): 161～168
[27] Welsh J, McClelland M. Fingerprinting genomes using PCR with arbitrary primers[J]. Nucleic Acids Research, 1990, 18:7213～7218
[28] Crowhust R N, Hawthorne B T, Rikkerink E H A, et al.Differentiation of Fusarium solani f.sp. cucurbitae race 1 and 2 by random amplification of polymorphic DNA[J].Curr.Genet.1991, (20): 391～396
[29] Assigbestse K B, Fernandez D, Doubios M P, et al. Differentiation of Fusarium oxysporium fsp. vasinfectum races on cotton by random amplified polymorphic DNA (RAPD) analysis[J]. Phytopathology, 1994, (84): 622～626
[30] 陈万权．冯洁，秦庆明．DNA分子标记在植物真菌病害研究中的应用．植物保护学报，1999，26(23)：277～282
[31] Marlatt M L, Correll J C, Kaufmann P, et al. Two genetically distance populations of fusarium oxysprum f. sp.lycopersici race3 in United states[J].Plant disease, 1996, 80(12): 1336～1342
[32] Nicholson P，Kezanoor H Z，苏海．用RAPD分析和DNA指纹鉴别玉米小斑病菌O、C、T三个小种[J]．植物病理学报，(23)：114
[33] Treena Burgess, Michael J, Brenda W. Simple Sequence Repeat Markers Distinguish among Morphotypes of Sphaeropsis sapinea. Applied and Environmental Microbiology, 2001, 67(1): 354～362
[34] Barve M P, Haware M P, Sainani M N, et al. Potential of microsatellites to distinguish four races of Fusarium oxysporum fisp.ciceri prevalent in India. Theor Appl Genet,
2001, 102: 138～147
[35] Elias K S, Zamir D, Lichtman-Pleban T, et al. Population structure of fusarium oxysporum f.sp.l ycopersicil: RFLP provide genetic evidence that vegetative compatibility groups is an indicator of evolutionary origin[J]. MPMI, 1993, 6(5): 565～572
[36] Manicom B Q, Baayen R P. Restraction fragment length polymorphisms in fusarium oxysporum f.sp. dianthi and other fusaria from dianthus species[J],Plant Pathology, 1993, 42:851～857
[37] Okoli C A N, Carder J H, Barbara D J. Restriction fragment length polymorphisms(RFLPs)and there relationships of some host-adapted isolates of Verticillium dahliae[J]. Plant Pathology, 1994, 43: 33～40
[38] Cander J H, Barbara D J. Molecular variation and restriction fragment length polymorphic (RFLPs) within and between six species of Verticillium[J]. Mycological research, 1991, 95: 935～942
[39] Boman E S, Khush G S, Nelson R J. Genetic differentiation among isolates of pyricularia grisea infecting rice and weed hosts [J]. Photopathology, 1993, 83: 393～399
[40] Milgroom M G, Lipari S E, Enno R A, et al. Comparison of genetic diversity in the chestnut blitht fungus, Cryphonectria parasitiea, from China and the U.S [J].Mycological research, 1992, 96: 114～120
[41] Vainio E J, Korhonen K, Hantula J. Genetic variation in Phlebtopsis glgantecn as detected with random amplified mierosatellite (RAMS) markers[J], Mycol Res, 1998, 102(2): 187～192.
[42] Paul W Tooley, Nichole R O'Neill, Erin D Goley, et al. assessment of diversity in Claviceps africana and other Claviceps species by RAM and AFLP analyses[J]. Phytopathology, 2000, 90(10): 1126～1130.
[43] Hantula J, Müller M M. Variation within Gremmeniella abietina in Finland and other countries as determined by Random Amplified Microsatellites (RAMS)[J]. Mycol. Res.1997, 101(2): 169～175
[44] 陈洪，朱立煌，李冬梅，等．致病性念珠菌DNA的AFLP指纹图谱[J]．科学通报，1996，41(10)：935～938
[45] Majer D, Mithen R, Lewis B G, et al. The use of AFLP fingerprinting for the detection of genetic variation in fungi[J]. Mycol Res, 1996, 100(9): 1107～1111
[46] Baayen R P, O' Donnell K, Bonants P J M, et al. Gene Geneal: ogies and AFLP Analyses in the Fusarium oxysporum Complex Identify monophyletic and nonmonophyletic formae speciales causing wilt and rot disease[J]. Phytopathology, 2000, 90(8): 891～900
[47] Folkertsma R T, Vanzandvoot P M, De Groot K E, et al. Gene pool similarities of potato cyst nematode populations assessed by AFLP analysis[J]. Mol Plant-Microbe Interact, 1996, (9): 47～54
[48] Kaplsn M, Cas well-Chen E P, Lliamson V M. Assessment of host-induced selection on three geographic isolates of Heterodera schachtii using RAPD and AFLP marker[J]. Phtopathology, 1999, 89:68～73
[49] Agus P. Genetic diversity of isolates of the leptosphaeria raaeulans species complex from Australia, Europe and North America using amplified fragment length polymorphism analysis[J].Mycological Research, 2000, 104(7): 772～781
[50] 邹亚飞．棉花黄萎病菌(Verticillium dahliae)致病型AFLP分析与分子鉴定研究[D]．重庆：西南农业大学，2001
[51] 高俊莲，陈文新，Terefe W Z，等．应用AFLP技术对斜茎黄氏根瘤菌遗传多样性的分析研究[J]．应用与生物学报，1999，5(4)：387～395
[52] 何秋月，Hei Leung，Zeigler Robert S，等．稻瘟病菌变异株的AFLP分析[J]．菌物系统，2002，21(3)：363～369
[53] Namikli F, Shiomi T, Nishi K, et al. Pathogenic and genetic in the Japanese strains of fusarium oxysporum f.sp melons[J], phatopathology, 1998, 88(8): 804～810
[54] Welsh J. Fingerprinting genomes using PCR with arbitrary primers[J]. Nucleic Acids Research, 1990, (18): 7213～7218
[55] Williams G K. DNA polymorphisms amplified by arbitrary primers are useful as genetic markers[J]. Nucleic Acids Researvh, 1990, (18): 6531～6535
[56] 马峙英，王省芬，张桂寅，等．现代生物技术在棉花黄萎病菌分化研究中的应用[J]．河北农业大学学报，1998，21(2)：102～111
[57] Crowhust R N. Differentiation of Fusarium solani fisp. cucurbitae race 1 and 2 by random amplification of polymorphic DNA[J].Curr.Genet, 1991, 20:391～396
[58] 陈万权．DNA分子标记在植物真菌病害研究中的应用[J]．植物保护学报，1999，26(3)：277～282
[59] Femandez D, Tantaoui A. Random amplified polymorphic DNA(RAPD) analysis: A tool for rapid characterization of Fusarium oxysporum f.sp.albedinis isolates[J]. PhytopatholMediterr, 1994, 33: 223～229
[60] Bentley S, Pegg K G, Dale J L. Genetic.variation among a world-wide collection of isolates of Fusarium oxysporum f.sp cubense analysed by RAPD-PCR fingerprinting[J]. Myeol.Res, 1995, 99:1378～1384
[61] Manulis S, Kogan N, Reuven M, et al. Use of the RAPD technique for identification of Fusarium oxysporumf.sp dianthi from carnation[J]. Phytopatholigy, 1994, 84:98～101
[62] Mes J J, Van Doom J, Roebroeek E J A, et al. Detection and identificatuib of Fusarium oxysporum fsp. gladioli by RFLP and RAPD analysis[J]. Modern Assays for Plant Pathogenic Fungi, 1995, 63～68
[63] Grajal-Martin M J, Simon C J, Muehlbauer F J, Use of random amplified polymorphic DNA (RAPD) to characterize race 2 of Fusarium oxysporum fsp. pisi[J]. Phytopatholigy, 1993, 83:612～614
[64] Zabeau M, Vos P. Selective restriction fragment amplification: a general method for DNA fingerprinting[J]. European Patent Application. 1993,
[65] 李珊，赵桂仿．AFLP分子标记及其应用[J]．西北植物学报，2003，23(5)：830～836
[66] 王斌，翁曼丽．AFLP的原理及其应用[J]．杂交水稻，1996，(5)：27～30
[67] 杜金友，黎裕，王天宇，等．SSR和AFLP分析玉米遗传多样性的研究[J]．华北农学报，2003，18(1)：59～63
[68] 易干军，谭卫萍，霍合强，等．龙眼品种(系)遗传多样性及亲缘关系的AFLP分析[J]．园艺学报，2003，30(3)：272～276
[69] 宗绪晓．D Vaughan．A Ka发a．等．AFLP分析小豆(Vigna angularis)种内遗传多样性[J]．作物学报，2003，29(4)：562～568
[70] 朱申龙，Morti M L，Rao R．应用AFLP方法研究中国大豆的遗传多样性[J]．浙江农业学报，1998，10(6)：302～309
[71] 贾建航，李传友，金德敏，等．AFLP技术在玉米自交系类群分析研究中的应用[J]．高技术通讯，1999，(4)：43～47
[72] 高世斌，李晚忱，荣廷昭，等．玉米骨干自交系AFLP指纹图谱鉴定[J]．四川农业大学学报，2001，19(2)：126～128
[73] 罗安定，陈守才，吴坤鑫，等．AFLP在橡胶树优异种质研究中的应用[J]．植物学报，2001，43(9)：941～947
[74] 马渐新，周荣华，董玉琛，等．小麦抗条锈病基因定位及分子标记研究进展[J]．生物技术通报，1999，(1)：1～6
[75] 黄祖六，谭学林，Tragoonrung．S．等．稻米直链淀粉含量基因座位的分子标记定位[J]．作物学报，2000，26(6)：777～782
[76] 房经贵，刘大钧，马正强．利用双杂合位点标记资料构建芒果遗传图谱[J]．分子植物育种，2003，1(3)：313～319
[77] 张蕴哲，刘红霞，邬荣领，等．毛新杨×毛白杨AFLP分子遗传图谱[J]．林业科学研究，2003，16(5)：595-603
[78] 康国斌，林德佩，王叶筠，等．西瓜枯萎病菌镰刀菌酸对西瓜苗作用机制的初步探讨[J]．植物病理学报，2000，30(4)：373～374
[79] 王浩波，王鸣，马德伟．用生物间遗传学原理研究西瓜与枯萎病菌的相互关系[J]．河北农业大学学报，1994，17(3)：29～33
[80] Kim W K. Isolation of high molecular weight DNA and double -stranded RNAs from fungi[J].Canadian Journal of Botany,1990, (68): 1898～1902
[81] Rogers S O.Extraction of DNA from Basidiomycetes for ribosomal DNA hybridizations[J].Canadian Journal of Botany, 1989, (67): 1235～1243
[82] 吴少慧，张成刚，张忠泽．RAPD技术在微生物生物多样鉴定中的应用[J]．微生物学杂志，2000，20(2)：44～47
[83] Paterson A H, Brnbaker C L, Wendel J F. A rapid method for extraction of cotton(Gossypium spp.) genomic DNA suitable for RFLP or PCR analysis[J]. Plant
Molecular Biology Reporter, 1993, 11(2): 122～127
[84] Van Toai, Martin S. Optimization of silver-staining AFLP technique for soybean[J]. SoybeanGenet, Newsl, 1996, (23): 206～209
[85] 田清震，盖钧镒，喻德跃，等．大豆DNA扩增片段长度多态性(AFLP)研究[J]．大豆科学，2000，19(3)：210～217