2个抗虫棉的外源Bt基因分子鉴定及其染色体定位

以中美2个抗虫棉品种GK19与33B为试验材料,利用检测中美Bt基因的特异性引物,分别对抗虫棉亲本GK19和33B进行PCR扩增,并通过SSR分子标记技术对其Bt基因进行分子鉴定与染色体定位,旨在从外源基因转化事件的视角探究中美转基因抗虫棉差异的分子基础。结果表明, GK19为中国转Bt基因抗虫棉, 33B为美国转Bt基因抗虫棉; GK19的Bt基因被定位在棉花Chr.20上,共16对SSR多态性标记与其Bt基因连锁,两侧的分子标记为NAU3907和NAU2579,其遗传距离分别为2.4 cM和1.5 cM; 33B的Bt基因被定位在棉花Chr.26上,共20对SSR多态性标记与Bt基因连锁,目标Bt基因位于标记NAU460和dc40260之间,其遗传距离分别为3.6 cM和2.0 cM。以上结果表明GK19和33B属于不同的遗传转化事件。     

中国金针菇工厂化生产用种SSR和AFLP遗传多样性分析

探讨SSR、AFLP分子遗传标记技术在金针菇遗传变异分析和菌种鉴定上的应用。用SSR和AFLP分子标记技术比较分析了中国市场上20个主要的工厂化生产金针菇用种的遗传多样性。29对SSR引物和60对AFLP引物分别检测到37个及206个多态位点;遗传相似系数分别为0.56~1.00、0.54~0.97。AFLP标记的多态性比SSR高。2种标记的聚类分析结果均揭示出中国市场上工厂化金针菇用种遗传多样性相对较小,亲缘关系十分接近。     

萝卜抽薹基因连锁的AFLP和SCAR分子标记鉴定

本文运用AFLP分子标记技术,结合BSA法,对萝卜抽薹基因相连锁的AFLP分子标记进行研究,获得两个与萝卜耐抽薹基因相连锁的AFLP标记,遗传距离分别为14.6 cM和9.1 cM.序列测定和Blast分析表明,标记ACG-CAG(123 bp)与拟南芥中锌指蛋白3(zinc finger protein 3,ZFP3)的cDNA同源性为89%,期望值为6e-23;标记ACT-CTG(175 bp)与拟南芥中的Copia类反转录转座子AtRE1基因的同源性为83%,期望值为4e-42.并将其中一个标记转化为简单易行的SCAR标记,遗传距离为7.5 cM.     

小麦抗白粉病新基因的AFLP和SSR标记及其染色体定位

M53 (YAV2/TEZ//Ae.squarrosa 249) 是硬粒小麦与粗山羊草的双二倍体合成种,携带一个抗白粉病新基因,暂命名为Pm-M53,该基因对北京地区白粉病优势生理小种15号表现免疫抗性。本研究利用来源于杂交组合M53/宛7107的一个F₂群体,在苗期采用白粉病15号小种（Blumeria graminis f. sp. tritici）接种,抗病反应型鉴定表明,抗感比例符合3∶1,说明其抗性受显性单基因控制;对部分F₂植株的F₃株系的抗病鉴定进一步证明了F₂鉴定的可靠性;利用AFLP和SSR标记技术结合F₂分离群体对目的基因进行了遗传作图,将目的基因定位在5D染色体的长臂上。其中AFLP标记P16M16_-109(Apm109)和P5M16_-161(Apm161)与目的基因的遗传距离分别为1.0和3.0 cM。SSR标记Xwmc289b、Xgwm583和Xgwm292与目的基因的遗传距离分别为20.0、33.0和24.0 cM。这些标记位于目的基因的两侧。利用中国春遗传背景的缺-四体和双端体结合AFLP标记Apm109确证了SSR标记定位的可靠性,进一步证明该基因是一个新的抗白粉病基因。     

甘蓝型油菜细胞质雄性不育恢复基因的SSR标记及初步定位

以甘蓝型油菜恢复系Q 16C和不育系2116A为亲本构建F2群体,根据BSA法的构建原则,使用F2群体分别构建了可育基因池与不育基因池。运用98对SSR引物对油菜亲本及基因池进行了多态性分析,结果表明,有1对引物(OS 31)能在亲本和基因池间表现出多态性,使用该引物继续对F2群体的230个单株进行分析,表明标记(OS 31)与恢复基因Rfp的遗传距离为0.08cM,可作为恢复系辅助育种的候选标记。     

水稻显性早熟基因Ehd的SSR标记定位

以籼稻品种广陆矮4和粳稻品种台中65为亲本构建高世代回交分离群体,选用分布于水稻全基因组的145个SSR标记对亲本及抽穗期早熟基因进行分析.结果表明,114个标记在亲本间具有多态性,多态率78.6%;在BC3F1群体中,检测到10个标记的基因型来源于供体亲本广陆矮4号;在BC3F2定位群体中,早熟植株数与晚熟植株数的分离比例为3:1,早熟植株平均比晚熟植株提早抽穗21 d;通过SSR标记与抽穗期共分离分析将显性早熟基因Ehd界定在分子标记RM271和RM258之间;Ehd与标记RM184和RM271紧密连锁,遗传距离分别为2.6 cM和2.1 cM,此结果为该基因分子标记辅助选择奠定了基础.     

黄瓜远缘群体分子遗传连锁图谱的构建和分析

以野生黄瓜品种和普通栽培黄瓜品种作亲本获得的142个F2群体为材料,采用AFLP,SRAP,SSR等分子标记进行遗传分析,构建了包含10个连锁群,有159个标记组成的黄瓜遗传连锁图谱,其中包括112个AFLP标记,39个SRAP标记和8个SSR标记。该遗传图谱覆盖基因组长度743.11 cM,平均图距4.67 cM。     

水稻特异亲和基因S-e的分子定位

水稻籼粳亚种间杂种具有强大的优势，但亚种间杂种的不育性限制了这一优势的利用。开展杂种不育基因的定位工作，对于进一步了解杂种不育性的遗传基础，克服亚种间杂种的不育性具有重要的意义。本研究选用粳型品种台中65的近等基因系E47-1和籼型品种广陆矮4号为材料，利用74个SSR标记对杂种F2群体进行偏态分离标记的筛选，同时根据F2和F3群体花粉育性和具有偏态分离的SSR标记之间的连锁关系，对特异亲和基因(F1花粉不育基因)S-e座位进行了分子定位，取得了以下主要结果：1、利用116个均匀分布在水稻12条染色体上的SSR标记对籼粳两亲本进行多态性筛选。结果有101个SSR标记在亲本间具有多态性，15个SSR标记在亲本间无多态性，SSR标记在亲本间的多态率高达87．07％。2、选用74个亲本间具有多态性的SSR标记对E47-1／广陆矮4号组合F2群体的偏态分离进行了初步的筛选和分析。发现有6个染色体区段的9个SSR标记在F2群体中存在偏态分离，它们分别位于第3、第6、第7、第10、第11和第12染色体上，卡方值均达到显著或极显著水平。6个染色体区段中有2个严重偏态分离区段，分别位于第6和第12染色体。3、通过对F2群体的花粉育性和偏态分离区段的SSR标记基因型的相关关系分析，表明位于第12染色体上的SSR标记RMl9附近存在一个F1花粉不育基因。继而在该标记附近设计位置特异性微卫星标记PSM401、PSMl80、PSMl82，利用F3作图群体，将特异亲和基因S-e座位定位在分子标记PSM401、PSMl80和PSMl82、RMl9之间，该基因与各标记的遗传距离分别为2．3cM、1．3cM、3．7cM和4．3cM。4、选取在S-a、s-b、S-c、S-d、S-e五个座位均纯合、花粉表现为部分不育的F2单株，发展了另一R群体，表明该群体存在另一特异亲和基因座s-f。本研究利用SSR标记，对特异亲和基因S-e进行了分子定位。S-e座位的分子定位，进一步丰富和完善了特异亲和性的学术观点，并为分子标记辅助选育水稻的粳型亲籼系奠定了基础。     

梨AFLP分子连锁图谱的构建与分析

遗传图谱是数量性状基因定位、基因图位克隆及分子辅助育种等研究的基础,而目前国内在梨树上这方面的研究尚属空白.试验以早美酥×八月红的杂交F1 92株实生苗为作图群体,采用了扩增稳定、多态性丰富的16对Mse I/EcoR I引物组合,对分离群体进行AFLP多态性检测,共获得208条多态性带,其中偏离孟德尔遗传的比率29.8%(P<0.01).应用Joinmap 3.0软件且结合"双假测交"策略,对208个AFLP分子标记进行了遗传连锁分析,构建了一张包含145个AFLP标记,分属20个连锁群的梨分子遗传连锁图谱,每个连锁群包含4～20个标记,平均为9.06个标记,图谱总长度覆盖梨基因组618.7 cM,标记间平均图距为6.58 cM.此图谱为梨树基因定位、比较基因组学和重要农艺性状的QTL定位等研究奠定了基础.     

与茄子青枯病抗性相关基因连锁的AFLP标记研究

本文以茄子高感青枯病亲本自交系5810、高抗亲本自交系5556单1、两自交系杂交得到F1,F1单株自交得到F2群体为试材,采用苗期接种鉴定及BSA法结合AFLP技术,探讨了茄子青枯病抗性的遗传规律,获得与茄子青枯病抗性相关基因连锁的AFLP分子标记。结果表明：抗病亲本5556单1青枯病抗性受一对单隐性基因控制,感病亲本控制的感病基因是不完全显性的。在抗、感池中未发现标记,而通过抗、感池单株分析得到了相引相AFLP分子标记E13M10,如及相斥相AFLP标记E16M5240,估算它们与目标基因间的遗传距离分别为10．14cM和7．56cM。     

Molecular mapping of a new gene for resistance to frogeye leaf spot of soya bean in 'Peking'

Amplified fragment length polymorphism (AFLP) and microsatellite (simple sequence repeat, SSR) techniques were used to map the _RGSp_eking gene, which is resistant to most isolates of Cercospora sojina in the soya bean cultivar ‘Peking’. The mapping was conducted using a defined F₂ population derived from the cross of ‘Peking’(resistant) בLee’(susceptible). Of 64 EcoRI and MseI primer combinations, 30 produced polymorphisms between the two parents. The F₂ population, consisting of 116 individuals, was screened with the 30 AFLP primer pairs and three mapped SSR markers to detect markers possibly linked to Rcs_Peking. One AFLP marker amplified by primer pair E‐AAC/M‐CTA and one SSR marker Satt244 were identified to be linked to Res_Peking. The gene was located within a 2.1‐cM interval between markers AACCTA178 and Satt244, 1.1 cM from Satt244 and 1.0 cM from AACCTA178. Since the SSR markers Satt244 and Satt431 have been mapped to molecular linkage group (LG) J of soya bean, the Res_Peking resistance gene was putatively located on the LG J. This will provide soya bean breeders an opportunity to use these markers for marker‐assisted selection for frogeye leaf spot resistance in soya bean.     

甘蓝型油菜白花性状的遗传及AFLP标记

甘蓝型油菜的白花性状是一种有用的指示性状,本研究以甘蓝型白花油菜及其分离群体为材料,对甘蓝型油菜的白花性状的遗传规律进行了研究。结果表明,甘蓝型油菜的白花性状受1对不完全显性基因控制。利用集团分离法(BSA)对白花基因进行AFLP分析,在256对AFLP引物中共获得6个与白花基因紧密连锁的分子标记。EA11MC04与EA12MC01为基因两侧最近的分子标记,其遗传距离分别为0.8,1.0 cM。     

Identification of AFLP markers linked to the epistatic suppressor gene of a recessive genic male sterility in rapeseed and conversion to SCAR markers

A recessive epistatic genic male sterility (REGMS) two‐type line, 9012AB, has been used for rapeseed hybrid seed production in China. The male sterility of 9012AB is controlled by two recessive duplicate sterile genes (ms1 and ms2) interacting with one recessive epistatic suppressor gene (esp). Homozygosity at the esp locus (espesp) suppresses the expression of the recessive male sterility trait in homozygous ms1ms1ms2 ms2 plants. In this study, we used a combination of bulked segregant analyses and amplified fragment length polymorphism (AFLP) to identify markers linked to the suppressor gene in a BC₁ population. From the survey of 1024 AFLP primer combinations, eight markers tightly linked to the target gene were identified. The two closest markers flanking both sides of Esp, P9M5₃₇₀ and S16M14₇₈₀, had a genetic distance of 1.4 cM and 2.1 cM, respectively. The AFLP fragment from P4M8₁₉₀, which co‐segregated with the target gene was converted into a sequence characterized amplified region marker. The availability of linked molecular markers will facilitate the utilization of REGMS in hybrid breeding in Brassica napus.     

芥菜型油菜多室基因Bjmc2的精细定位

芥菜型多室油菜的产量比普通两室油菜更高,定位乃至克隆多室基因可为油菜遗传改良及解释多室角果形成机制创造条件。本研究通过验证JD11-2家系衍生群体仅在BjMc2位点上存在差异,可用于BjMc2的定位。采用AFLP结合BSA法分析BC₅和BC₆群体,筛选到1个与BjMc2连锁的AFLP标记并转化为SCAR标记SC1。基于该AFLP标记序列信息,利用白菜同源序列设计SSR引物和SCAR引物,获得11对SSR标记和1对SCAR标记。通过在芥菜型油菜BAC文库中的挑选,获得2个覆盖目标区域的单克隆,由此开发1个SSR标记。将获得的SCAR和SSR标记扫描BC₇群体,构建了两室性状基因BjMc2的遗传连锁图,两侧最近标记ZX17和BACsr96与目标基因之间的遗传距离分别为0.048 cM和0.340 cM,并定位到白菜A7 scaffold000019的946~1014 kb之间,约68 kb物理距离。     

Genetic mapping for the Rf1 (fertility restoration) gene in sunflower (Helianthus annuus L.) by SSR and TRAP markers

The Rf1 gene in sunflower can effectively restore the pollen fertility of PET1 cytoplasm in male-sterile lines and has been widely used in commercial hybrid production. Identifying molecular markers tightly linked to this gene will be useful in marker-assisted selection to develop maintainer and restorer lines. Rf1 has been mapped to Linkage Group (LG) 13 of the public sunflower simple sequence repeat (SSR) map by aligning maps constructed from different populations and only one SSR marker was reported to be loosely linked to Rf1 . This paper reports the result of applying target region amplification polymorphism (TRAP) and SSR markers to map and develop a sequence-tagged site (STS) marker tightly linked to Rf1 using two populations derived from a cross between two U.S. public sunflower lines, RHA439 and cmsHA441. An SSR marker, ORS511, was 3.7 cM from the Rf1 gene and a TRAP marker, K11F05Sa12-160, was linked to Rf1 at a distance of 0.4 cM. This TRAP marker was converted to an STS marker for using in sunflower breeding.     

A genetic linkage map of rhodesgrass based on an F1 pseudo-testcross population

The linkage relationships between 164 polymorphic amplified fragment length polymorphism (AFLP) and 25 restriction fragment length polymorphism (RFLP) fragments assayed in a pseudo‐testcross population generated from the mating of single genotypes from two divergent cultivars were used to construct female, ‘Katambora’ (‘Kat’) and male, ‘Tochirakukei’ (‘Toch’) parental genetic maps for rhodesgrass. The ‘Kat’ genetic map consists of 84 marker loci (72 AFLP and 12 RFLP markers) distributed on 14 linkage groups and spans a total length of 488.3 cM, with an average distance of 7.8 cM between adjacent markers. The ‘Toch’ genetic map consists of 61 marker loci (52 AFLP and nine RFLP) mapped on 12 linkage groups spanning a total length of 443.3 cM, with an average spacing of 9.0 cM between adjacent markers. About 23% of the markers remained unassigned. The level of segregation distortion observed in this cross was 11.1%. In both maps, linked duplicated RFLP loci were found. These linkage maps will serve as a starting point for linkage studies in rhodesgrass with potential application for marker‐assisted selection in breeding programmes.     

Development of molecular markers linked to the wheat powdery mildew resistance gene Pm4b and marker validation for molecular breeding

Powdery mildew, caused by Blumeria graminis (DC.) E.O. Speer f. sp. tritici, is an important disease in wheat (Triticum aestivum L.). Bulk segregant analysis (BSA) was employed to identify SRAP (sequence‐related amplified polymorphism), sequence tagged site (STS) and simple sequence repeat (SSR) markers linked to the Pm4b gene, which confers good resistance to powdery mildew in wheat. Out of 240 SRAP primer combinations tested, primer combinations Me8/Em7 and Me12/Em7 yielded 220‐bp and 205‐bp band, respectively, each of them associated with Pm4b. STS‐₂₄₁ also linked to Pm4b with a genetic distance of 4.9 cM. Among the eight SSR markers located on wheat chromosome 2AL, Xgwm382 was found to be polymorphic and linked to Pm4b with a genetic distance of 11.8 cM. Further analysis was carried out using the four markers to investigate marker validation for marker‐assisted selection (MAS). The results showed that a combination of the linked markers STS_?241, Me8/Em7_?220 and Xgwm382 could be used for marker‐assisted selection of the resistance gene Pm4b in wheat breeding programmes.     

21种不同类型葡萄种质资源遗传多样性的SSR分析

本研究通过SSR标记解析不同类型葡萄种质遗传多样性差异和亲缘关系,为资源分类与品种鉴定提供理论依据。利用15对SSR引物对21种不同类型葡萄种质进行PCR扩增,对扩增情况、遗传距离与亲缘关系等进行分析。15对SSR引物共扩增出等位基因位点91个,其中多态性位点89个,多态性比率高达97.8%,其中13对引物扩增多态性百分率达到100%。21份供试材料间遗传相似系数变化范围在0.45~0.91之间,并在遗传相似系数0.62处被分为2个类群。研究表明SSR标记从分子水平上解析了不同类型葡萄种质的遗传差异性,揭示了材料间亲缘关系,为育种研究提供了一定参考依据。     

小麦合成种M53抗白粉病基因的RAPD和SSR标记

运用RAPD和SSR技术，采用分离群体分组分析法(BSA)进行了小麦合成种M53抗白粉病基因连锁的分子标记研究。结果表明，M53的抗白粉病基因由显性单基因控制，RAPD标记OPL09-1700与抗病基因连锁，遗传距离为16.8cM。SSR标记Xgwm205也与抗白粉病基因连锁，遗传距离为9.3cM，通过SSR标记将该基因定位于5DS，标记与基因间的排列顺序     

利用陆海杂种BC1群体构建遗传连锁图谱并初步定位产量性状相关的QTL

摘要：本研究利用中棉所36和海1配制杂交组合,并用中棉所36为轮回亲本构建回交群体(BC1F1, BC2F1和 BC1S1)。用亲本和F1对新开发的2102对SSR引物进行多态性筛选,共筛选到317对含有海1显性带的引物,占筛选引物总数的15.08%;最终对其中的275对引物进行了BC1F1群体扩增,获得306个SSR标记差异位点。连锁分析表明（LOD=6.5）,有254个标记位点连锁,分布在42个连锁群中,覆盖2252.36cM,约占棉花基因组的50.05%;平均每个连锁群有6.08个标记,覆盖53.63 cM;标记间平均间距为8.87cM。利用BC1F1、BC2F1和BC1S1三个不同世代分离群体产量性状数据,共定位16个产量性状QTL,解释表型变异5.77%～19.86%。其中,衣分6个,铃重6个,籽指4个。有9个增效基因来自陆地棉亲本中棉所36,7个增效基因来自海岛棉亲本海1,说明了表型性状较差的品种同样可能含有可用于性状改良的增效基因。控制衣分的3个QTL可在不同的世代稳定检测到,效应稳定,增效基因均来自高值亲本陆地棉,为进一步分子标记辅助选择奠定了基础。