基于KASP技术的小麦条锈菌SNP分子标记开发与评价

为开发用于小麦条锈菌Puccinia striiformis f. sp. tritici群体遗传研究的竞争性等位基因特异性PCR-单核苷酸多态性(kompetitive allele specific PCR-single nucleotide polymorphism,KASPSNP)标记,以中国小麦条锈菌流行小种CYR32的基因组为参考,与美国小麦条锈菌流行小种PST78和印度小麦条锈菌流行小种38S102的基因组进行比对,根据比对到的SNP位点设计KASP-SNP引物,用64个中国小麦条锈菌标样对其进行筛选,同时用13对多态性引物组成的简单重复序列(sim‐ple sequence repeat,SSR)分子标记分析这64个标样,并利用Powermarker 3.25和Structure 2.3软件通过多态性指数和群体遗传结构分析来评价KASP-SNP和SSR两种分子标记。结果显示,共比对到29 929个SNP位点,设计出462对KASP-SNP引物,经64个中国小麦条锈菌标样筛选到43对多态性较好的引物,所开发的这43对KASP-SNP引物多态性信息含量指数平均为0.346,基因多样性指数平均为0.420,而SSR引物的2种指数分别为0.237和0.265,前者较后者分别高出46.0%和58.5%。2种标记结果的群体遗传结构分析可得到类似结果,最佳聚类数K值都为4,云南菌系是遗传结构相对最简单的菌系,湖北菌系是遗传结构相对最复杂的菌系,但个别菌株的遗传划分存在较大差异。表明本研究开发的KASP-SNP分子标记多态性较SSR分子标记更加丰富,具有较好的应用前景。     

小麦抗源Sw92抗叶锈病基因遗传及其分子标记

以小麦优异抗源Sw92为父本,感病小麦品种铭贤169为母本,杂交获得F1、F2和BC1代群体。采用我国叶锈菌优势小种PHT对双亲及其杂交世代进行接种鉴定。结果表明,小麦抗源Sw92对叶锈菌小种PHT的抗性系由一对隐性基因所控制。采用简单重复序列(SSR)技术对Sw92携带的抗性基因进行分子标记,共筛选了371对SSR引物,获得2个引物(WMC494、WMC737)可在抗/感池和双亲中扩增出多态性DNA片段。遗传连锁分析结果表明,该抗病基因位于小麦6BS上,与WMC494、WMC737标记的遗传距离分别为3.4cM和15.0cM,不同于6BS上的已知抗叶锈基因Lr36和Lr53,暂命名为LrSw92。     

100个小麦品种资源抗条锈性鉴定及重要抗条锈病基因的SSR检测

了解小麦品种资源对中国条锈菌生理小种的抗病性水平及其所含重要抗条锈病基因,可为合理种植和利用小麦品种提供理论依据。选用中国小麦条锈菌不同生理小种CYR17、CYR32、CYR33和V26及条锈菌混合小种分别对100个小麦品种资源进行苗期和成株期抗条锈性鉴定,并采用SSR分子标记技术检测重要抗条锈病基因Yr5、Yr10、Yr15、Yr18、Yr24和Yr26。结果表明,在苗期对4个生理小种均表现抗病性的有‘Mos311’、‘兰天15号’等30个品种;在成株期表现抗病性的有‘Yeoman’、‘兰天1号’、‘小红麦’等88个品种;苗期和成株期均抗病的有‘Mos311’、‘兰天15号’等30个品种。SSR检测发现,‘贵农22’可能含有Yr5,‘兰天23号’、‘兰天24’号、‘中梁04260’和‘中梁04335’可能含有Yr10,‘Little Joss’和‘中梁5号’可能含有Yr15,‘清农3号’、‘兰天2号’、‘中梁04335’等19个品种可能含有Yr18,检测品种可能均不含Yr24和Yr26。在鉴定中保持稳定抗病性的‘Mos311’、‘兰天15号’等8个品种在抗病品种选育中有重要应用价值。     

引进ICARDA的小麦苗期抗旱性鉴定及SNP关联分析

以引进国际干旱地区农业研究中心（ICARDA）的160份小麦为研究对象,在苗期用20% PEG-6000模拟干旱条件进行处理,以正常营养液作为对照,分析干旱环境对7个苗期相关性状（苗高、最长根长、根数、茎叶鲜质量、茎叶干质量、根鲜质量和根干质量）的影响,利用综合评价值进行抗旱评价及抗旱分级,并结合55K SNP芯片对159份小麦的苗期抗旱相关性状的抗旱系数进行关联分析。抗旱鉴定结果表明：干旱处理下的各个苗期相关性状均低于正常处理水平,按照综合评价值及系统聚类,将160份小麦分为高抗旱（5份）、中等抗旱（53份）、低抗旱（97份）和干旱敏感（5份）四类;在苗期筛选出5份高抗旱品种,包括ICARDA69、ICARDA51、ICARDA49、ICARDA83、ICARDA84,其D值分别为0.823、0.813、0.765、0.722、0.711。关联分析结果显示：利用24151个SNP标记位点结合苗期相关性状的抗旱系数在P≤0.001水平下共定位到227个抗旱相关标记,分布在除1B、2D、4D和6D外的17条染色体上,可解释7.13%~14.68%的表型变异。检测到3个多效应位点,分别位于4B、5B和6A染色体上,可解释9.31%~13.28%的表型变异。其中,位于4B染色体上的AX-108789337与茎叶干质量和根干质量显著关联,可解释10.44%~13.28%的表型变异;位于5B染色体上的AX-109353092与苗高和根鲜质量显著关联,可解释9.31%~10.93%的表型变异;位于6A染色体上的AX-110432128与苗高和根鲜质量显著关联,可解释9.95%~9.99%的表型变异。     

SSR marker analysis of Monilinia fructicola from Swiss apricots suggests introduction of the pathogen from neighbouring countries and the United States

Monilinia fructicola is a quarantine fungal pathogen in Europe, but many major stone fruit growing countries in Europe have reported its presence recently. In Switzerland, the fungus was first found in a single apricot orchard in 2008. This study confirms the presence of M. fructicola in nine out of 22 commercial orchards in Canton Valais, Switzerland. Five simple sequence repeat markers (SSRs) were developed for M. fructicola and samples from Switzerland, Spain, Italy, France and the United States were analysed and compared in order to assess the genetic diversity of the pathogen, identify the origin of the disease, and verify if the fungus reproduces sexually in Europe. In the 119 European samples analysed, 12 different haplotypes were found, indicating a relatively high genetic diversity of the pathogen considering that the first report in Europe was 10 years ago. Three haplotypes found in Europe were identical to those found in the American samples (two from the east coast and one from the west coast). Population structure analysis suggests that the European population is derived from at least two ‘invasion’ events probably originating from the US (one from the east coast, the other from the west coast). Preliminary evidence of sexual reproduction of M. fructicola in Europe is reported.     

大豆疫霉多态性SSR标记开发及遗传多样性分析

 用FastPCR软件在大豆疫霉全基因组中搜索到1 234个含2~4个重复基元精确SSRs。选择260个SSRs设计引物,经对大豆疫霉5个分离物的基因组DNA检测,有212对(81.5%)有效扩增出SSR特征条带,112对(52.8%)扩增多态性。用18对多态性SSR引物分析了来自美国、中国黑龙江省和福建省大豆疫霉分离物的遗传多样性,在73个分离物中共扩增出112个等位变异,变异范围为4~9,平均为6.22个,表明选择的引物对具有高的多态性。在3个大豆疫霉群体中,黑龙江省和福建省分离物的遗传距离最近,美国和福建省分离物的遗传距离最远。UPGMA聚类将73个分离物划分为6组,其中8个美国分离物(72.73%)和53个中国分离物(85.48%)被聚类在一起,表明大豆疫霉中国分离物与美国分离物可能具有共同的祖先,中国分离物可能为外来种。     

Genetic analysis of Grapevine leafroll‐associated virus 3 population from Galicia,Spain

Grapevine leafroll‐associated virus 3 (GLRaV‐3; Ampelovirus, Closteroviridae) isolates from Galicia in northwestern Spain were selected to characterize their genetic diversity according to different factors (age, origin, location, variety, etc.). The vines belonged either to local white and red varieties autochthonous to Galicia or to varieties from other Spanish regions but widely used in Galicia. These GLRaV‐3 isolates came from different vineyards in Galicia located in coastal or inner areas. Multiplex RT‐PCR allowed the detection of isolates belonging to groups I, II, III–V and VI. Two genomic regions were studied in the isolates, the HSP70h and the capsid protein, using specific primers that allow the detection of variants from groups I to V. Some possible recombinants could be detected; however, multiple infections with different variants indicated that they were not genuine recombinants. No differences were found in the population structure considering variety or geographical factors. Isolates belonging to four groups were found in the distinct areas surveyed: groups I and II were the most common, followed by groups VI and III, as is the case in the rest of the world. In the same surveys, the presence of insect vectors for GLRaV‐3 was investigated and found lacking in inland areas but present in those with milder climate. Genetic analysis did not support isolation of the GLRaV‐3 isolates in Galicia, suggesting that the uncontrolled exchange of infected vines and/or rootstocks has been a major agent of virus spread.     

基于分子遗传标记技术的舞毒蛾种群遗传关系研究进展

在简要介绍以形态、行为、生态学为依据的舞毒蛾分类发展的基础上,概述了分子遗传标记技术在舞毒蛾种群鉴定、起源、分化、入侵和扩散方面的研究进展,初步分析了我国舞毒蛾种群遗传关系研究的现存问题,并提出建议。     

25份北欧小麦抗白粉性评价及抗白粉病基因标记鉴定

采用苗期人工接菌的方法鉴定了25份北欧小麦对河南省7个不同地区小麦白粉病菌分离物的抗性,同时利用与已知抗白粉病基因Pm2、Pm3f、Pm4、Pm6、Pm13、Pm16、Pm21和Pm24连锁的分子标记对其中抗性优良的材料进行基因检测。结果表明,8份材料对所有接种的白粉病菌菌株表现高抗或免疫;其中NGB7476和NGB7809携带Pm4a和Pm16基因, NGB7481和NGB9954携带Pm6和Pm16基因,NGB9955和NGB9956同时携带Pm4a、Pm6和Pm16基因,8份材料均不携带Pm2、Pm3f、Pm4b、Pm13、Pm21 和Pm24基因。