黑木耳单核杂交菌株遗传分析

为实现黑木耳菌株快速鉴定和栽培性状定向育种,研究通过荧光核染色筛选,酯酶同工酶酶谱比较及RAPD分子鉴定,筛选出了黑木耳杂交单核体的交配核型及杂交新种,并对杂交新种进行了遗传分析和栽培比较.结果显示,2个亲本筛选出4种不同类型原生质体单核体,8808亲本单核体为A1、A2型,981亲本单核体为B1、B2型.单核体单-单杂交获得核型为A1B2的杂交菌株,命名为Z1.酯酶同工酶谱及RAPD图谱结果显示Z1菌株含有来源单核的互补型条带和亲本条带.聚类分析将被试菌株分为3类,Z1菌株与981相似度高于8808菌株.栽培试验显示Z1与981栽培性状更近似而又集合了双亲的性状优势.研究的开展可为将来黑木耳菌株鉴定及杂交子遗传分析等提供快速、可靠检测方法.     

运用交配型基因分子标记辅助蛹虫草亲和配对

为了建立一种分子标记辅助育种技术体系以增加蛹虫草杂交亲和性配对的比例,以4个蛹虫草菌株(GD1、JS2、SD2、中13)为供试亲本,通过无性孢子分离方法获得了8株单交配型的杂交亲本,根据蛹虫草交配型位点MAT-alpha和MAT-HMG中基因序列设计了分子标记并指导配制亲和性组合,最终成功获得了12个杂交菌株。在栽培试验的基础上,分析了12个杂交菌株的栽培周期、产量、活性成分含量(虫草素和腺苷)等相关性状,结果表明,蛹虫草杂交菌株存在超亲现象。本研究建立的这套以交配型基因作为分子标记辅助选育蛹虫草新品种的高效杂交育种技术体系,将有助于促进蛹虫草栽培产业的发展。     

用ISSR分析四川苎麻品种(系)间的遗传关系及雄性不育分子标记的建立

本文从100条ISSR引物中筛选出21条引物,对8个四川苎麻品种(系)间的遗传关系进行了分析。PCR扩增结果表明,21条引物在8份材料中共扩增出86条带,平均每条引物扩增出4.1条,其中多态性位点71个,各引物扩增出的位点数3~8个不等,平均每条引物可以检测到3.4个多态性位点。聚类分析和遗传距离分析结果表明,供试品种川7、川8与其亲本遗传距离较远。3个杂交品种(川7、川8、川9)均表现特有的偏父本遗传现象。此外,本研究用ISSR引物U835筛选到了1个雄性不育分子标记,并将其扩增产物克隆测序,结果表明该序列大小为658bp。根据该序列,此标记被转化成稳定的SCAR标记,可用于苎麻雄性不育分子标记辅助育种。     

我国“应县小黑豆”对SCN4号生理小种抗性的SSR及ISSR分子标记研究

本研究以"应县小黑豆"×"晋豆23"杂交组合F2群体为材料,用塑膜钵柱法进行抗性鉴定,利用分离体分组分析法(BSA)结合SSR和ISSR标记技术对抗性基因进行分子标记筛选和定位,旨在实现大豆孢囊线虫抗性育种中亲本和杂交后代的分子标记辅助选择.筛选到2个与抗SCN基因座位连锁的分子标记Satt038和ISSR811.Satt038片段含180bp和185 bp,为共显性标记,在F2群体中的分离比为121;ISSR标记ISSR811为显性标记,片段长350bp,F2群体分离比为13.2个标记在F2群体中呈孟德尔式遗传.通过JOINMAP分析,微卫星标记Satt038、ISSR标记ISSR811与抗SCN基因座位的遗传距离分别为26.9cM和10.2 cM.还探讨了分子标记Satt038和ISSR811用于大豆抗SCN育种进行分子标记辅助选择的可能性.     

多花黑麦草杂交种SSR分子标记鉴定及表型比较分析

多花黑麦草为异花受粉植物,杂交过程中易出现杂交种生物性混杂现象,有必要对杂交后代进行真假鉴定分析.本研究旨在筛选出多花黑麦草优势杂交组合和优异杂交后代材料,为多花黑麦草杂交育种工作提供基础资料.实验利用SSR分子标记辅助鉴定5个多花黑麦草(Lolium multiflorum)骨干品种为亲本创制的杂种后代真实性,并对亲本与杂种后代主要表型进行比较.结果表明:(1)用20对引物组合共得到多态性条带426条,多态性位点百分率为82.41％,说明SSR分子标记能够很好的揭示多花黑麦草材料间的差异;(2)经SSR分子鉴定,将后代分为有父本特征谱带的杂种后代、无父本特征带的后代2类,在150个杂交后代单株中有父本特征谱带的杂种后代有108个为真杂种,其中杂交组合GC、CG真杂种最多均为24个,WG组合真杂种最少为19个;(3)杂交组合CG的F1代部分形态特征显著优于亲本,5个杂交组合内(间)在营养器官与生殖器官性状均存在显著差异,其中杂交组合MG的变异系数最高,WG的变异系数最低,杂交组合CG形态特征显著高于其他杂交组合.由研究结果可以初步判断长江2号×赣选一号多花黑麦草为优势杂交组合,其杂交种为优异杂交后代材料,可为后续杂交选育新品系提供后备材料.     

茶树种质资源ISSR分子标记初步研究

本研究采用100个ISSR引物对32个茶树资源的DNA进行PCR扩增反应,其中UBC826、UBC847、UBC849U、BC850、UBC854、UBC860、UBC873、UBC881共8个引物能扩增出较好的多态性,根据引物UBC854、UBC881的多态性能将32个茶树资源进行区分,初步建立32个茶树资源的ISSR分子指纹图谱。研究表明ISSR能有效用于茶树种质资源鉴定,茶树ISSR反应具有较高的稳定性。     

杂交水稻亲本分子身份证及SSR指纹数据库的建立

本研究利用GB/T20396-2006推荐的48对SSR核心标记,对133份杂交水稻亲本(包括部分常规品种)进行SSR分析,根据引物在133份材料中扩增的片段大小,构建了各品种的分子身份证和指纹图谱,该身份证由48位数字或数字与字母组合组成,代表了48对引物在该品种中的扩增情况。将品种的身份证信息与品种的形态学特征特性等信息相结合,构建了具查询功能的SSR指纹网络数据库。该SSR指纹数据库除具有品种基本信息查询、品种分子身份证查询、品种间相似率查询和身份证核对等基本功能外,还可指导杂交水稻品种真实性和纯度鉴定,分析亲本间遗传相似性,为杂交稻新组合的选育提供亲本选配的参考。该数据库的建立,不仅为检测单位和育种工作者提供了一个便利的网络数据交换和共享平台,也为品种的选育、试验、管理、评价、仲裁提供技术支撑和科学参考。     

利用目标起始密码子多态性(SCoT)分子标记分析梨遗传多样性

目标起始密码子多态性(start codon targeted polymorphism,SCo T)是一种新型的遗传分子标记技术。本研究采用单因素水平设计方法,优化并建立梨的适宜SCo T分子标记体系,通过SCo T标记技术,分析了安徽省砀山县的43份梨(Pyrus spp.)的遗传多样性和亲缘关系。结果表明,优化后梨SCo T-PCR反应体系:10×Easy Taq Buffer(含2 mmol/L Mg2+)2.5μL,模板DNA终浓度30 mg/L,上下游引物终浓度1.0μmol/L,d NTPs终浓度0.2 mmol/L,Taq DNA聚合酶1.0 U,总体积为25μL。随机选取2个DNA模板进行引物筛选,最终从67条引物中筛选出16条扩增条带清晰且有差异性的引物,共扩增出145条带,其中多态性条带有126条,多态性比率为86.1%,每条引物平均扩增出9.1条带。SCo T标记的43个梨材料遗传相似性系数为0.517~0.931,平均值为0.685。聚类分析表明,在遗传相似系数为0.610的水平上,43份梨材料分为A和B两组;在遗传相似系数为0.746的水平上,A组又分为6个亚组(Ⅰ~Ⅵ)。所研究的43份梨材料具有丰富的遗传多样性,且能够被SCo T分子标记有效地检验,为进一步研究利用安徽省砀山县梨种质资源提供了依据。     

猪基因组AFLP指纹图谱的构建

摘要：对猪（Sus scrofa）基因组AFLP指纹图谱构建的各项条件进行了优化,包括双酶切消化、接头连接、预扩增和选择性扩增、凝胶电泳银染技术及荧光标记检测法等,建立了稳定的猪基因组DNA的AFLP指纹图谱构建方法,采用E32/T32引物组合在45个猪种基因组混合DNA中检测到28个多态标记。     

SSR和SRAP标记研究油菜杂交种骨干亲本的遗传多样性

用SSR和SRAP两种分子标记方法研究51份甘蓝型油菜杂交种亲本系的遗传多样性,并对两种分子标记研究结果进行比较。结果发现,在51份材料中,45对SSR引物共扩增出194条多态性条带,平均每对引物为4.3条,25对SRAP引物共扩增出197条多态性条带,平均每对引物为7.9条。UPGMA聚类分析表明,SSR和SRAP标记都可将51份亲本材料划分为五大类群,本所选育的玻里马细胞质雄性不育系（Polima CMS）的主要保持系和恢复系都聚在同一类群的不同亚群中。根据系谱资料分析发现,SRAP标记划分的类群与系谱资料更为接近,SRAP标记更适用于遗传关系较近材料的遗传多样性分析。SRAP标记揭示的亲本间遗传距离要大于SSR标记揭示的遗传距离。两种不同标记方法揭示出油菜亲本遗传多样性的差异主要是由不同的标记方法揭示的标记位点等位基因变异数不同造成的。     

辐照花粉辅助授粉对百合远缘杂交结实的影响及杂种早期分子鉴定

研究了辐照花粉辅助授粉对百合‘pollyanna’(♀)×毛百合杂交结实的影响。结果表明:采用普通柱头授粉能获得许多膨大果实和饱满种子,表明该组合为亲合性组合。用经1000和2500Gy的60Coγ射线辐照的母本花粉进行辅助授粉明显提高了果实中饱满种子的结籽率,表明2种剂量的辐照花粉辅助授粉均能有效克服该组合的杂交障碍。结籽数的提高能获得更多的后代,增加了从F1代中选育新优单株的几率。随机选取辐照花粉辅助授粉获得的杂交子代15株,采用ISSR技术对其组培苗进行亲子鉴定,6条ISSR随机引物在15个子代中共扩增出372个条带,其中具有父本特征带114条,具有母本特征带67条,具有双亲共有带65条。ISSR鉴定结果证实了杂种的真实性。     

RAPD and ISSR molecular markers in <Emphasis Type="Italic">Olea europaea</Emphasis> L.: Genetic variability and molecular cultivar identification

Thirty Portuguese and eight foreign olive (Olea europaea L.) cultivars were screened using Random Amplified Polymorphic DNA (RAPD) and Inter-Simple Sequence Repeat (ISSR) markers. Twenty RAPD primers amplified 301 reproducible bands of which 262 were polymorphic; and 17 ISSR primers amplified 204 bands of which 180 were polymorphic. The percentage of polymorphic bands detected by ISSR and RAPD was similar (88 and 87%, respectively). The genetic variability observed was similar in the Portuguese and foreign olive cultivars. Seven ISSR and 12 RAPD primers were able to distinguish individually all 38 olive cultivars. Twenty specific molecular markers are now available to be converted into Sequence Characterised Amplified Region (SCAR) markers. Relationships among Portuguese and foreign cultivars is discussed.     

Genetic diversity and relationships among safflower (<Emphasis Type="Italic">Carthamus tinctorius</Emphasis> L.) analyzed by inter-simple sequence repeats (ISSRs)

Genetic diversity and relationships among 48 safflower accessions were evaluated using 22 inter-simple sequence repeats (ISSR) primers. A total of 429 bands were amplified, and 355 bands (about 82.7%) were polymorphic. Five to forty-one polymorphic bands could be amplified by each primer, with an average of 16.1 polymorphic bands per primer. The results showed that the polymorphism of the safflower germplasm was higher at the DNA level. All the 48 accessions could be distinguished by ISSR markers and were divided into 9 groups based on ISSR GS by using UPGMA method. The genetic relationships among the accessions from different continents were closer. Comparatively, the genetic diversity of the accessions originated from Asia was higher, from Europe assembled. The results also showed that the genetic variation of accessions from Indian and Middle Eastern safflower diversity centers were relatively higher. ISSR is an effective and promising marker system for detecting genetic diversity among safflower and give some useful information on its phylogenic relationships.     

利用ISSR标记分析福云（半）同胞系茶树品种（系）的遗传多样性和亲缘关系

福鼎大白茶和云南大叶茶是茶树育种中的骨干亲本,利用这两个亲本选育了许多优良的品种（系）。本研究利用ISSR标记技术分析了40个福云（半）同胞系茶树品种（系）的遗传变异水平和亲缘关系。14个ISSR引物在供试品种（系）间共扩增出251条谱带,多态性条带的比率为96.4％。引物的PIC值平均为0.94,Rp值平均为29.35,表明引物扩增位点的高多态性和对品种的强辨别能力。40份供试品种（系）的基因多样性指数（H）为0.35,Shannon信息指数（I）为0.52。按母本来源和育种机构对供试品种（系）进行分组分析,结果表明不同品种（系）组间的遗传多样性水平比较接近,遗传变异主要存在于组内品种（系）的个体之间,不同组间基因交流明显。供试品种（系）间的相似系数介于0.52-0.75,根据相似系数矩阵按UPGMA法对40个供试品种（系）进行聚类分析,构建了不同品种（系）间的亲缘关系树状图。福鼎大白茶在树状图中形成单独的分支,在树状图的根基处,其它品种（系）根据遗传距离聚类成不同的类群。来源于同一育种单位的部分茶树品种（系）聚类在同一类群中,但未发现按母本来源区分的独立类群。总之,通过ISSR标记分析,可在基因组水平上进一步了解福云（半）同胞系茶树品种（系）间的遗传变异水平,并进一步明确其亲缘关系,为今后福云（半）同胞系在茶树育种上的有效利用提供依据。     

Assessment of genetic diversity in Solanum trilobatum L., an important medicinal plant from South India using RAPD and ISSR markers

Solanum trilobatum L. is an Indian medicinal plant containing rich amount of steroidal glyco-alkoloids that can be used as precursor for commercial steroid production. Two efficient marker systems such as Random Amplified Polymorphic DNA (RAPD) and Inter Simple Sequence Repeats (ISSR) were used for the first time to assess the genetic diversity across 14 S. trilobatum accessions obtained from five South Indian states. Twenty out of 60 RAPD primers generated 189 distinct bands of which 160 were polymorphic with an average of 8 polymorphic bands per primer. A maximum of up to 15 fragments were amplified with an average of 9.45 bands per primer and the amplicons varied in size between 100 and 3,000 bp. The percentage of polymorphism ranged from 55.5 to 100, with an average of 84.6. ISSR profiling using 7 out of 20 primers amplified 83 bands and the number of amplified fragments varied from 2 to 16 with a size range of 200–1,800 bp. Totally 72 polymorphic bands were obtained using 7 ISSR primers at an average of 10.28 polymorphic bands per primer. Polymorphism percentage varied from 50 to 100 among the selected accessions resulting in an average percentage of polymorphism of 86.7. The PIC values ranged from 0.49 to 0.93 for RAPD and 0.16 to 0.90 for ISSR primers. The study pointed out that ISSR markers were more efficient than RAPD markers in evaluating the degree of genetic variation in S. trilobatum. The UPGMA cluster analysis grouped all Tamil Nadu accessions in one cluster and other state accessions in another cluster. The Principal component analysis also substantiates this clustering pattern. Thus the phylogenetic relationship and a high genetic variation revealed in the present study could provide a baseline data for conservation and improvement of this plant in future. Also the molecular markers identified in this study will be helpful in authentication of this species to prevent adulteration in herbal medicine.     

2个马铃薯杂交组合F1的SSR鉴定

杂交育种是马铃薯新品种选育的重要方法, 其杂种F₁真实性鉴定是获得目标性状单株的关键环节。为选育优质、高产、抗病性及抗旱性强的马铃薯新品种, 用马铃薯品种"费乌瑞它"(Favorita)分别与"J07-6"和"陇薯3号"杂交, 获得了杂种F1代。本试验利用SSR标记技术对"Favorita"×"J07-6"、"Favorita"×"陇薯3号"2个杂交组合F1共86个单株的真实性进行了鉴定。试验从43对SSR引物中筛选出2对适宜引物STM1049和S7。利用这2对引物进行PCR扩增, 将"Favorita"×"J07-6"杂交种F1和"Favorita"×"陇薯3号"杂交种F1的SSR带型划分为双亲互补型、缺失型、父本型和母本型4种类型。依据SSR带型特征, 从"Favorita"×"J07-6"和"Favorita"×"陇薯3号"2个杂交组合F₁单株中分别鉴定出真杂种34个和27个, SSR分子标记技术用于马铃薯杂交种真实性鉴定是可靠的。该研究结果可为马铃薯杂交种优良株系选育提供依据。     

斑鳢、乌鳢及其杂交种遗传差异的AFLP分析

本文采用AFLP分子标记技术对斑鳢、乌鳢和杂交鳢（斑鳢（母本）与乌鳢（父本））共85个个体（其中斑鳢、乌鳢各30个,杂交鳢25个）进行了遗传差异分析。结果表明11对引物组合共检出了459个不同的扩增片段,扩增出的多态谱带数350条,多态性比例为76.25%,平均每对引物组合扩增出31.8条多态条带。乌鳢与斑鳢种群间存在稳定的、可以简单地借以进行群体鉴别的标记条带169条,其中父本（乌鳢）特异性条带78条,72条能够稳定地遗传给杂交鳢;母本（斑鳢）特异性条带89条,71条能够稳定地遗传给杂交鳢。杂交鳢另外出现了3条非双亲的条带。遗传差异的分子方差分析结果发现,斑鳢与乌鳢种群间的遗传相似度为0.5161,杂交鳢与斑鳢和乌鳢种群间的遗传相似度相近,分别为0.7189和0.7476,斑鳢与乌鳢之间以及杂交鳢与斑鳢和乌鳢之间的种群间遗传距离分别为0.6615、0.3300和0.2909,即AMOVA分析显示斑鳢、乌鳢和杂交鳢间存在极显著的遗传分化。UPGMA聚类分析显示,在个体间,斑鳢与乌鳢能区分成两大类,杂交鳢则分散于斑鳢和乌鳢种群中;在群体间,杂交鳢首先与乌鳢聚类,然后与斑鳢相聚,表明杂交鳢种群总体上更偏向于父本乌鳢。研究结果表明,杂交鳢与斑鳢和乌鳢发生混杂的可能性很大,应该对杂交鳢进行隔离养殖。本文结果将为斑鳢、乌鳢和杂交鳢的遗传分析提供实验依据,也为其种质的合理利用提供参考。     

Analysis of genetic diversity of Ruthenia Medic (Medicago ruthenica (L.) Trautv.) in Inner Mongolia using ISSR and SSR markers

Ruthenia Medic is tolerant to drought, cold, high salinity, resistance to trampling and high quality features. Inter-simple sequence repeat (ISSR) and simple sequence repeat (SSR) molecular markers were employed for the first time to access the genetic diversity and relationships of 30 wild Ruthenia Medic accessions obtained from Inner Mongolia in the present study. A total of 94 bands were amplified by ten ISSR primers, of which 83 (88.5 %) were polymorphic, and 57 polymorphic bands (80.4 %) were observed in 69 bands amplified by ten SSR primers. Shannon’s information index (I = 0.487), and average expected heterozygosis (He = 0.329) generated by ISSR primer were higher than that of SSR analysis (I = 0.372, He = 0.231). The study indicated that ISSR were more effective than SSR markers for assessing the degree of genetic variation of Ruthenia Medic. UPGMA cluster analysis revealed inconsistencies in the clustering patterns, as the Mantel’s test between the dendrograms for ISSR and SSR data indicated a poor fit for the ISSR and SSR data types (r = 0.0970). Whereas the pattern of clustering of the genotypes remained relatively the same in ISSR and combined data of ISSR and SSR. The results of principal components analysis also supports their UPGMA clustering. These results have an important implication for Ruthenia Medic germplasm characterization, improvement, and conservation.