稻纹枯菌酯酶同工酶、可溶性蛋白及致病力的研究初报

 对稻纹枯菌25个菌株分别进行了致病力测定、酯酶同工酶和可溶性蛋白分析.结果表明:不同菌株存在明显的致病力分化,而酯酶同工酶图谱却有较强的一致性,它们的主酶带基本相同,但在副酶带上反映出一定的同工酶表型异质性,表现为带的数目及着色深浅的不同;可溶性蛋白图谱比酯酶同工酶图谱表现出更多的多样性,大多数致病力较强的菌株都有l~2条特征谱带,而且它们的谱带数目比弱毒力菌株的多出3~5条.     

稻瘟病菌生理小种酯酶同工酶特性的研究

 运用聚丙烯酰胺凝胶等电聚焦法对来自江、浙两省经全国统一鉴别寄主鉴定的稻瘟病菌7群12个生理小种的16个菌株菌体酯酶同工酶进行了分析比较,结果表明:稻瘟菌菌体酯酶同工酶带有13-18条,等电点分布范围为3.00-7.20。在等电点4.15附近有各生理小种共同的酶带。同一生理小种群内不同生理小种间酯酶同工酶的相似系数一般大于0.50,不同生理小种群的生理小种间酶谱的相似系数一般为0.30-0.40,也存在少数不是同一生理小种群的小种间酶谱相似系数大于0.50的情况。供试各生理小种间酶谱均存在不同程度的差异,表现为酶带数目的多少和等电点分布的不同。来自不同省份经鉴别寄主鉴定为同一生理小种的菌株间酶谱的相似系数亦仅0.05-0.60。研究结果初步反映出供试大多数稻瘟菌菌株间菌体酯酶同工酶的相似性与致病性有关,但也受菌株来源地生态环境影响。本文还讨论了稻瘟病菌致病性容易变异但在许多情况下又具有相对稳定性的可能机制,探讨在一定范围内聚丙烯酰胺凝胶等电聚焦技术用于稻瘟菌生理分化和致病性变异监测的可能性及尚待进一步研究解决的穗题等。     

新疆野生啤酒花群体遗传多样性的RAPD分析

用RAPD技术分析新疆7个啤酒花(Humulus lupulus L.)的天然居群遗传多样性及群体间的遗传分化,12个随机引物共检测到121个可重复的位点,其中多态位点119个,占总位点的98.35%.由Shannon表型多样性指数和Nei基因多样性指数估计,居群内遗传分化百分比分别为63.55%和65.65%,表明啤酒花种内的遗传变异主要存在于群体内.分析认为,新疆境内啤酒花自然居群的遗传多样性很丰富,居群间产生了一定的分化.啤酒花居群的基因流Nm=1.0037,相对有限的基因流可能在居群遗传分化的维持中起着作用.     

捕食线虫真菌酯酶同工酶谱和可溶性蛋白质谱分析

 本文对5个属14个种共33株捕食线虫真菌采用聚丙烯酰胺凝胶电泳技术进行了酯酶同工酶谱和可溶性蛋白质谱分析。结果表明,酯酶同工酶谱带少,但各属的特征谱带是明显的,在同属不同种之间,多数种都有自已的特征谱带与其它种相区别,同种不同菌株间没有明显差异;蛋白质谱带较多,在每个属中均有1~2条特征谱带,属间的谱带差异明显,同属不同种和同种不同菌株间蛋白质谱带都表现出多型性。     

赖草发育过程中同工酶的研究

利用聚丙烯酰胺凝胶电泳技术 ,对赖草酯酶和过氧化物酶同工酶研究结果表明 :赖草不同发育期的酯酶和过氧化物酶同工酶酶谱及酶活性存在明显的器官间差异和多态性 ;其中过氧化物同工酶与赖草生长发育的关系比酯酶相对密切 ,同时稳定性也较强 ,可以作为赖草生长发育特性的生理指标和遗传鉴定指标     

四川省不同来源核盘菌菌株的可溶性蛋白和酯酶同工酶的电泳图谱分析

选取四川省不同地区、不同寄主、代表不同致病性水平和不同菌丝体亲和性表现的核盘菌(Sclerotinia sclerotiorum)菌株进行可溶性蛋白和酯酶同工酶电泳分析,结果表明:不同寄主来源、不同致病性水平、不同菌丝体亲和性表现的核盘菌菌株间的可溶性蛋白和酯酶同工酶电泳图谱条带数量及分布相对稳定,仅部份弱带在出现频率、染色强度或迁移距离上有一定的变化,表现出种的特征和种内的异质性,在分类上具有一定的意义,可溶性蛋白和酯酶同工酶电泳分析可作为核盘菌分类和遗传多样性分析的一个生化指标。     

稻瘟菌不同小种间可溶性蛋白及酯酶同工酶的多态性

采用聚丙烯酰胺凝胶泳对来自太湖稻区的１０个水稻稻瘟病菌株和２个禾本科杂草瘟菌的可溶性蛋白及酯酶同工酶进行了分析比较。结果表明：供试菌株菌体可溶性蛋白可呈现１６～２５条电泳谱带，其中在Ｒｆ值为０．２３，０．３３，０．４４的３条谱带为１２２个菌株所共有，没有发现生理小种特异性谱带。对１２个菌株α－酯酶同工酶的分析表明，在所有菌株同工酶谱带中可分辩出具有不同迁移率的１２条谱带，据此可将菌株分为８个不同的同工酶谱带类型。同时表明，α－酯酶同工酶在太湖稻区稻瘟病菌中存在丰富的多态性，但其谱带类型与病菌生理小种无相关性。     

胡杨叶形变化与POD同工酶的关系研究

以5个不同发育阶段的胡杨植株为研究对象,调查各发育阶段胡杨植株树冠不同层次当年生枝同一节位叶片的叶形指数,同时测定其POD同工酶酶谱和酶活性,探讨不同发育阶段胡杨植株叶形的变化规律和POD同工酶酶谱及酶活性间的关系。结果表明:(1)不同发育阶段的胡杨植株都表现出叶形指数随冠层的升高而减小;(2)不同发育阶段的胡杨植株都表现出单株不同冠层叶片中POD同工酶表达具有高度的稳定性和相似性,酶谱带型基本一致,而不同发育阶段的胡杨植株间POD同工酶酶谱在质(酶带数目和迁移率)和量(酶的活性)上表现出不同程度的差异;(3)不同发育阶段的胡杨植株均表现为树冠不同层次当年生枝叶片的叶形指数与POD酶活性极显著正相关(r=0.668,p<0.01)。     

澳洲大蠊二种同工酶谱及胆碱酯酶活性的研究

本文采用聚丙烯酰胺凝胶盘状电泳研究澳洲大蠊三个发育阶段酯酶同工酶，过氧化物酶同工酶谱的变化，并分别测定了发育三阶段和成虫头、胸、腹、脚胆碱脂酶活性。结果发现过化物酶同工酶与酯酶同工酶酶谱各有特征，从卵到二龄幼虫发育过程中表现最大活性的同工酶的酶带位置各不相同，胆碱酯酶活性在卵、一龄、二龄幼虫中以一龄幼虫活性最高，成虫中以头部活性最高，腹部活性最小。     

十八种赤眼蜂同工酶的比较研究

本实验用聚丙烯酰胺垂直平板凝胶电泳分析了18种赤眼蜂的酯酶、超氧化物歧化酶和苹果酸酶同工酶。赤眼蜂的酯酶同工酶有明显的种间差异,超氧化物歧化酶同工酶也有一定的差异,苹果酸酶同工酶种间差异较小,但某些种有较独特的酶谱。     

稻曲病菌遗传多样性与群体结构的初步分析

 利用随机扩增多态性DNA (random amplified polymorphic DNA,RAPD)初步分析了稻曲病菌(Ustilaginoidea virens)的群体遗传结构。从1 60个随机引物中筛选32个扩增带型清晰、重复性好的引物,对不同年份采自辽宁、云南、湖北和浙江等水稻种植区的5 6个菌株进行扩增。32个引物扩增出2 2 3条带,绝大多数引物对不同年度采自不同稻区的菌株扩增的DNA谱型相同,大多数菌株间相似性系数达0.80以上。根据扩增DNA片段的多态性,从空间分布来看,来源于北方、长江流域和南方的菌株难以划分出明显的地理宗谱;不同年度的菌株DNA多态性也无明显的差异。上述结果初步表明稻曲病菌遗传稳定,寄主选择作用(寄主的基因型及其时空分布)对稻曲病菌变异的影响较小。但是尚需采用其它的分子技术测试更多的菌系,才能较系统地分析我国稻曲病菌系的遗传变异及群体结构特点。     

甘薯茎腐病菌的遗传多样性及致病力差异分析

为了解不同地区甘薯茎腐病菌Dickeya dadantii种群遗传多样性水平及致病力差异,采用重复序列PCR基因指纹(repetitive element palindromic PCR,REP-PCR)技术和薯片接种方法,对采自广东省、广西壮族自治区和重庆市的6个市区县的59株菌株进行分析。结果表明,5对引物对59株菌株扩增出41个清晰的条带,其中36个为多态性条带,每对引物的扩增条带数在4～10之间,平均为7.2。在物种水平上,有效等位基因数、Nei’s基因多样性指数和Shannon信息指数分别为1.4768、0.2801和0.4186,其中湛江种群多样性最高,南宁种群多样性最低;当遗传相似系数为0.79时,59株菌株可被划分为5个类群,类群划分与菌株来源地间有一定的相关性。此外,不同地区病菌种群间存在明显的致病力差异,其中合浦种群与湛江种群致病力最强,万州种群致病力较弱。表明甘薯茎腐病菌种群具有丰富的遗传多样性,不同地区的病菌种群存在明显的遗传多样性与致病力差异。     

Genetic diversity of the annual weed Monochoria vaginalis in southern China detected by random amplified polymorphic DNA and inter-simple sequence repeat analyses

Two molecular genetic screening techniques, RAPD (random amplified polymorphic DNAs) and ISSR (inter-simple sequence repeats), were applied to detect the level and pattern of genetic diversity of Monochoria vaginalis, a common weed of rice fields, in seven populations from southern China. Among these populations, 116 bands were amplified by 18 RAPD primers, of which 34 bands (29.31%) were polymorphic, and 14 ISSR primers produced 111 bands with 87 polymorphic bands (78.38%). Within each population, a relatively low level of genetic diversity was detected by both RAPD and ISSR analyses, with a mean genetic diversity (H) of 0.0348 and 0.0551 respectively. Analysis of molecular variance of the data from the RAPD and ISSR markers detected that the majority of total genetic variation existed among populations (73.50% and 76.70% respectively) and only minor genetic variation within populations (26.50% and 23.30% respectively). Cluster analysis divided the seven populations into two groups, indicating that the genetic relationships among populations have relatively low correlation with their geographical distribution (Mantel test; r = 0.45 and 0.48 respectively). Our results indicated that both RAPD and ISSR markers were effective and reliable for accurately assessing the degree of genetic variation of M. vaginalis. Comparing the two techniques, ISSR markers were more efficient than the RAPD assay. The Mantel test gave r = 0.16, suggesting no correlation between these two molecular markers.     

四川籼稻区稻瘟病菌群体遗传结构

应用rep-PCR分子指纹技术对2000～2002年采自四川6个籼稻自然生态区的137个稻瘟病菌菌株进行了DNA分子指纹扩增和聚类分析,共获得73个不同的DNA指纹图谱(单元型)和62条分子量不等的DNA带型.结果显示,无论以何种遗传相似水平划分,四川稻瘟病菌的群体结构都表现很突出的优势宗谱,又存在着具有较多遗传多样性的次要小宗谱和特异性宗谱,蕴含着极其丰富的遗传信息;在0.19遗传相似水平,所有供试菌株可以划分成37个遗传宗谱,层次较为丰富.四川稻瘟病菌群体结构具有明显的时空特点,不同年度间稻瘟病菌群体存在一定的亲缘关系,又各自拥有当年的特异性宗谱;在空间上,不同稻作区表现出从复杂到简单的病菌群体变化特点.稻瘟病菌的遗传宗谱与生理小种致病型不存在一一对应的关系,作者认为将二者横向比较没有可比性.     

Genetic diversity and differentiation of <Emphasis Type="Italic">Sclerotinia sclerotiorum</Emphasis> populations in sunflower

Ninety-six isolates of sunflower Sclerotinia sclerotiorum (Lib.) de Bary from Inner Mongolia (IM) in China, from Canada and the United Kingdom (UK) were sampled to investigate the genetic diversity and structure using Sequence-Related Amplified Polymorphism. A total of 123 polymorphic bands were obtained, ranging in size from 100 to 500 base pairs. The five populations of S. sclerotiorum isolated from the three countries showed various levels of genetic variability. The percentage of polymorphic loci varied from 30.89% in the UK population to 97.56% in the Middle IM population. The values of Shannon index (i) varied from 0.1876 in the UK population to 0.5301 in the West IM population. The heterozygosity of the five geographic populations obtained by estimating allele frequency varied from 12.91% in the UK population to 35.44% in the West IM population. The genetic identity, as indicated by the Nei unbiased identity index, ranged from 0.9744 between populations from Canada and East IM to 0.6477 between populations from West IM and UK. UPGMA cluster analysis using Nei’s genetic distance gave distances ranging from 0.0259 to 0.4343. The rates of gene flow among five geographic populations ranged from 1.5406 between West IM and UK populations to 18.4149 between West IM and Middle IM populations. The four populations from West IM, Middle IM, East IM and Canada were clustered into one subgroup in which the isolates from West and Middle IM belonged to one population, whereas those from East IM and Canada essentially were another population. The isolates from the UK formed a population that was significantly distinct from other populations.     

Molecular variability among isolates of <Emphasis Type="Italic">Mycosphaerella graminicola</Emphasis>, the causal agent of septoria tritici blotch,in Argentina

The genetic structure and diversity of Mycosphaerella graminicola population were studied with ISSR molecular markers, using isolates from several locations of the Argentinean wheat region: subregion IV (SE of Buenos Aires Province) and II South (central part of Buenos Aires Province). Samples were taken from different bread wheat (Triticum aestivum) cultivars. A total of 126 isolates were subjected to molecular analysis to compare the genetic structure of the isolates from both wheat subregions. Ten ISSR primers were used: (GACA)₄; (AAC)₇; (ATC)₇; (AC)₉; (AAG)₇; (AG)₉; (AGC)₅; (CAG)₅, (GTG)₅ and (GACAC)₃. Eighty-four bands ranging from 200 bp to 8.000 were amplified. Eighty-one distinct haplotypes were identified and 43 isolates did not generate any amplification products. The highest number of polymorphic DNA fragments were produced using ISSR primers (ATC)₇ and (GTG)₅, which detected bands in 38 isolates. The molecular analysis revealed the existence of 81 different haplotypes among the 126 isolates studied. These results revealed a high degree of genetic diversity in the M. graminicola population in Argentina.     

基于毒性相关基因序列的稻瘟病菌群体遗传多样性分析

 选用16对毒性相关基因特异性引物对四川和重庆9个县(市)分离到的200个稻瘟病菌单孢菌株进行PCR扩增,并采用最长距离法进行聚类分析,结果显示各引物均能扩增出其目的条带,多态位点百分率(P)高达93.75%,扩增频率差异较大;200个菌株可归为70个不同的单元型,其中单元型SCH13为优势单元型;在0.86遗传相似水平上,200个菌株可划分为27个遗传宗谱,包括1个优势宗谱,3个亚优势宗谱,14个次要宗谱,9个小宗谱,层次丰富;在群体平均水平上,病菌群体具有丰富的遗传多样性(H=0.324 4,I=0.484 2),且群体间差异较大;9个种群在遗传距离为0.05水平上可分为4个类群,种群遗传谱系与地理区域分布呈一定相关性。同时,该地区的群体存在一定的遗传分化(HT=0.320 0),群体内多样性大于群体间多样性(Hs=0.179 6,Dst=0.140 4),总遗传变异的56.13%存在于群体内(Gst=0.438 7),群体间基因流动性较小(Nm=0.639 6)。本研究揭示了四川和重庆部分区域稻瘟病菌群体遗传结构、遗传多样性及其与地理分布之间的关系,为抗病育种和品种布局奠定了基础。     

不同生境臭柏种群的遗传多样性分析及其与环境因子的相关性

毛乌素沙地臭柏群体是一个生态过渡带。为了进一步阐明分子变异和基因流与生境或生态过渡带的联系,应用RAPD标记开展了臭柏群体的分子生态学研究。采用随机扩增多态性DNA(RAPD)方法对臭柏(Sabina vulgaris.)的3个种群进行了研究.用11个随机引物扩增出129条清晰谱带,其中117条为多态性谱带。利用POPGENE3.2软件对数据进行处理,结果如下:(1)臭柏有着较丰富的遗传多态性,多态位点百分率达90.70%,各种群多态位点百分比在69.77%~72.87%之间.(2)臭柏的种群间分化较小Gst=0.1872,81.38%的遗传变异存在于种群内,各种群的遗传一致度都在86.22%.(3)聚类分析显示,生境相近的种群被聚到了一起,反映了臭柏种群的遗传分化和生境有着一定的相关性.又利用Nei,s指数统计了RAPD数据,也证实了大部分的遗传变异存在于群体之内。臭柏群体内的遗传多样性与土壤总钾呈显著的负相关。     

Genetic diversity and structure of the Fusarium oxysporum f.sp. lini populations on linseed (Linum usitatissimum) in China

The genetic diversity of specific Fusarium oxysporum f.sp. lini from six provinces in China was investigated using molecular markers, inter-simple sequence repeats (ISSR). Based on the morphological features and the internal transcribed spacer (ITS) sequences, 96 isolates were identified as Fusarium oxysporum. The 96 isolates were amplified by PCR with 12 ISSR primers. The number of bands amplified by each primer ranged from 43 to 142, with sizes ranging from 250 to 4,500 bp. A total of 800 bands were observed, out of which 797 were polymorphic (99.62%). The percentage of polymorphic loci varied from 17.25% in Gansu and Inner Mongolia to 33.75% in Sinkiang. Nei’s gene diversity index (h) ranged from 0.0428 in Gansu to 0.0666 in Sinkiang, and Shannon’s information index (I) ranged from 0.0675 in Gansu to 0.1117 in Sinkiang. The genetic identity using the Nei’s genetic identity varied from 0.9643 between the populations from Hebei and Gansu to 0.9844 between the populations from Sinkiang and Shanxi. Unweighted pair group mean analysis (UPGMA) cluster analysis, as indicated by the Nei’s genetic distance, showed the distances ranging from 0.0158 between the populations from Sinkiang and Shanxi to 0.0364 between the populations from Hebei and Gansu. The six populations were clustered into three subgroups. The Gansu population was clustered into one subgroup, the same as the Inner Mongolia population. The four other populations were clustered into the third subgroup. The Nei’s GST (0.2972) and gene flow among populations (Nm =1.1825) revealed large gene exchanges among populations.     

Extent and pattern of genetic differentiation within and between European populations of Phelipanche ramosa revealed by amplified fragment length polymorphism analysis

The extent and pattern of genetic differentiation between Phelipanche ramosa populations colonising tobacco in different European regions were investigated by amplified fragment length polymorphism (AFLP) analysis, in order to determine levels of variation for tobacco resistance breeding and management programmes. Four different AFLP primer pairs amplified a total of 1050 clear and reproducible bands, of which 962 (91.62%) were polymorphic among the 35 individuals taken from four P. ramosa populations collected in Spain, Italy, Bulgaria and Germany. Cluster analysis based on the AFLP data categorised the plants into distinct groups, in line with their geographical origin, denoting clear genetic differentiation among the four populations. This differentiation was supported by both high bootstrap values and significant results of the analysis of molecular variance. The most divergent population was the one from Bulgaria. The majority of the genetic diversity was attributable to differences among populations (77.80%), as expected from the predominant autogamous behaviour of this species. Populations differed significantly in within-population diversity, as measured by Shannon's information index. The German population presented the lowest genetic diversity and the Italian population harboured the highest level of within-population genetic diversity. There are significant differences in genetic diversity level among the studied P. ramosa populations and clear population-specific genetic diversity structures. These need to be taken into account, together with the potential differences in parasite aggressiveness, when planning breeding and management strategies for P. ramosa control in tobacco cultivation.