黄瓜无苦味基因bi的分子标记研究

以黄瓜纯合自交系9110Gt（bibi）和03828(BiBi)为双亲获得的F₂代为试材,利用分离群体组群分析法,从256对AFLP选择性引物中获得了与营养器官无苦味位点连锁距离为6.43 cM的AFLP分子标记TG/GCT₁₅₀,并成功地将其转化为操作简便、表现稳定的SCAR标记SC₈₇。     

白菜抗霜霉病基因的RAPD标记

研究了与抗霜霉病基因紧密连锁的RAPD分子标记, 利用白菜抗病品种014和感病品种010杂交的F₂群体144个单株为材料, 通过人工接种鉴定, 采用高抗单株和高感单株分别构建抗、感病池, 利用BSA法筛选了560条RAPD引物, 其中引物AY12在抗感病池中扩增出多态性片段AY12₁₂₃₈ , 通过F₂单株验证后证明AY12₁₂₃₈与抗霜霉病基因紧密连锁, 其遗传距离为6.7 cM。     

黄瓜遗传图谱构建及株高相关性状的QTL定位

 为利用分子标记辅助选择黄瓜矮生性状, 以矮生黄瓜D8 ×蔓生黄瓜J IN5的188株F^₂单株作为 作图群体, 应用ISSR和SRAP分子标记技术进行多态性筛选, 构建了含65个标记位点的遗传连锁图谱。整个图谱覆盖7个连锁群, 全长831.6 cM。标记平均间距为12.7 cM, 标记间最小遗传距离为4.8 cM, 最大遗传距离为22.3 cM。采用复合区间定位分析, 检测到控制黄瓜株高性状的QTL位点2个, 均位于第4连锁群上, 贡献率分别为13.2%和13.0%; 控制节间距的QTL位点1个, 也位于第4连锁群上; 控制第一雌花开花期的QTL位点7个, 分别位于第1、2、3、5、7连锁群上, 各QTL的贡献率在6.5%～12.5%之间。     

黄瓜全雌性基因连锁的AFLP和SCAR分子标记

 本研究以全雌品种‘戴多星’自交系和弱雌品种‘北京截头’自交系为双亲杂交获得F₁ ,然后得到F₂ 性型分离群体, 利用分离群体分组分析法(Bulked Segregant Analysis, BSA) 构建全雌和弱雌两个基因池, 筛选了64对AFLP选择性引物EcoR I-NN +Mse I-NNN组合, 发现EcoR I-TG +Mse I-CAC引物组合在全雌基因池中扩增出一条分子量为234 bp的特异带。经F₂ 代单株验证, 该特异条带能在全雌单株中稳定出现。以MAP MAKER (Version 310) 软件分析, 该标记与全雌性位点的连锁距离在617 cM。命名该连锁标记为TG/CAC₂₃₄。将该特异条带回收、克隆、测序, 设计特异SCAR引物, 再对F₂ 代单株基因组DNA进行扩增, 仅在全雌单株中扩增出1条分子量为166 bp 的特异带, 表明已成功地将与黄瓜全雌性连锁的AFLP标记转化为操作简便、表现稳定的SCAR标记, 该标记命名为SA₁₆₆。     

蔓生黄瓜与矮生黄瓜内源激素的动态变化

以蔓生黄瓜129和矮生黄瓜D0462为试验材料,采用酶联免疫吸附法（ELISA）测定了茎尖、叶片和根的内源激素GA、IAA、ZR和ABA含量,并对内源激素含量差异和动态变化进行比较分析。结果表明：矮生黄瓜D0462各器官的GA含量极显著低于蔓生黄瓜129;生长前期IAA含量高于蔓生黄瓜129,生长中后期IAA含量显著低于蔓生黄瓜129;ZR与ABA含量极显著高于蔓生黄瓜129。矮生黄瓜D0462各器官GA/ABA、IAA/ABA和（IAA+GA+ZR）/ABA的比值都极显著小于蔓生黄瓜129,而ZR/GA的比值极显著大于蔓生黄瓜129。由此推测,矮生黄瓜D0462矮化可能与内源GA含量低有关,并且矮化是多种激素共同作用的结果。     

南瓜矮生基因Bu的比较定位

 以中国南瓜矮生突变体为供体亲本,以印度蔓生南瓜为轮回亲本,构建了BC6F2分离群体。利用黄瓜基因组序列,将南瓜矮生基因Bu比较定位至黄瓜5号染色体,并开发了一个新的PCR标记IF3629,该标记与矮生基因Bu连锁遗传距离为1.0 cM。该标记不仅可以用于分子标记辅助选择育种,而且为Bu基因的克隆奠定了基础。     

金棚1号番茄种子纯度检测的SCAR标记

以金棚1号番茄及其父、母本为试材，提取叶片DNA，优化构建了稳定的RAPD反应体系。从在番茄上多态性丰富的100条随机引物中筛选获得1条可用于鉴别母本和F₁的引物S130，2条可用于鉴别父本和F₁的引物S296和S114。将扩增所得到的差异条带进行克隆、测序，设计出1对SCAR引物RF2/FF2，能特异鉴别F₁和母本。进一步建立了以干种子为试材检测金棚1号番茄种子纯度的SCAR标记检测体系。     

茄子单性结实基因的遗传分析及AFLP分子标记

以茄子单性结实自交系D-10和非单性结实自交系03-2为试验材料,研究了单性结实基因的遗传特性及其AFLP标记。对杂交后代F₁、F₂、BC₁单性结实性表现型分离比例的分析表明,茄子D-10的单性结实性由单显性核基因控制,其基因用符号Pat表示。采用AFLP分析技术和改良BAS法,通过512对E/M引物组合的筛选,获得1个与茄子单性结实基因紧密连锁的AFLP标记E75/M53-70,该标记与单性结实基因间的遗传图距为15.38 cM,可用于茄子单性结实性的鉴定和单性结实分子标记辅助育种,加速茄子单性结实基因的转育和利用。     

与黄瓜抗炭疽病相关基因连锁的AFLP标记的筛选

以黄瓜抗炭疽病母本66和感炭疽病父本A18及其F₂代分离群体为试材, 采用BSA法和AFLP技术建立了对炭疽病的抗病组和感病组, AFLP引物组合E24M48在抗感组间表现多态性, 且呈共显性。经220个F₂单株分析, 在高抗单株和高感单株中分别仅扩增出251 bp和245 bp的特异片段, 而在中间类型个体中同时扩增出了该两个特异片段。连锁值测定结果表明, 该特异带与黄瓜炭疽病抗病相关基因紧密连锁,距离为21727 cM。测序结果显示, 该两个片段的大小分别为251 bp和245 bp, 并且两个片段的差异在于两个“TCT”重复序列(6个碱基) 的插入或缺失。     

山葡萄种内和种间杂交后代果实成熟期遗传分析

 1973～2003年对我国东北山葡萄抗寒的种质资源(品种、品系) 进行种内杂交, 并与不抗寒的欧亚种酿造葡萄品种进行种间杂交、回交和重复杂交。共定植34个组合杂种苗, 成活5 639株。观察5种杂交模式, 34个组合5 639株杂种( F₁ , F₂ ) 苗的果实成熟期的遗传规律。5 种杂交模式果实成熟期(早→晚) 排列顺序是: 山葡萄×山葡萄≥ (山- 欧) F₁ ×山葡萄≥山葡萄×欧亚种≥ (山- 欧) F₁ ×(山- 欧) F₁ ≥ (山- 欧) F₁ ×欧亚种。遗传规律是: 山葡萄种内和种间杂交后代( F₁ , F₂ ) 果实成熟期分离, 表现为连续分布, 倾向于早熟亲本, 杂交组合中早熟亲本越多, 分离出的早熟单株越多, 为多基因控制的数量性状遗传、“早熟基因”有累加效应、呈显性遗传给后代。     

三角紫叶酢浆草叶色变异株系的RAPD和ISSR标记初步鉴定

 以三角紫叶酢浆草叶色变异株系和其野生型植株为材料,对其遗传物质进行分子水平上的初步检测。采用DNA混合样本池法,构建变异型样本池和野生型样本池,并以这两个DNA池做模板分别进行RAPD-PCR和ISSR-PCR 扩增。从70个RAPD随机引物和100个ISSR随机引物中分别筛选出2个RAPD特异引物（S76,S118）和2个ISSR特异引物（UBC818,UBC868）,均能在上述两个DNA池之间扩增出稳定的差异性条带共6条。其中,变异株系的差异性缺失带S118 700-800测序成功,其序列包含编码叶绿体光系统Ⅰ第Ⅶ亚基的基因。研究结果表明,与其野生型相比,该叶色变异株系的确发生了遗传物质的变化,且在一定继代范围内能通过无性繁殖方式稳定存在。     

Identification of RAPD markers linked to nut weight and plant stature in cashew

Cashew is an important edible nut crop of tropics. Bulk segregant analysis (BSA) was carried out on DNA bulks constituted from F₂ population and germplasm in order to link or associate molecular markers with economic characters. In all 458 RAPD, 31 ISSR and 21 pairs of SSR primers were used and identified polymorphic markers between parents. Though screening F₂ bulks with these markers identified markers polymorphic between the bulks but none could be validated with the individuals of their bulks. Hence screening with germplasm bulks was carried out and could identify four RAPD markers polymorphic between the bulks for nut weight and plant stature and also between the individuals of their bulks. Of the four, three markers were associated with nut weight amplifying at 775, 475, 275, bp region in primers OPN 14, UBC 184 and UBC 185 respectively. Out of these three, two markers were specific to low nut weight and one marker was specific to high nut weight and their bands were present in greater frequency (50–77.8% and 75%) of individuals constituting the respective bulks. Similarly, the another marker UBC 185₂₇₅ was detected which was specific to low plant stature and was present in 66.7% and 10% individuals constituting short and tall bulks respectively. Markers identified with bulks and with the individuals of bulks were validated further with more individuals of F₂ and germplasm.     

Breaking the intergeneric hybridization barrier in Carica papaya and Vasconcellea cauliflora

The present investigation was undertaken to develop PRSV (Papaya ringspot virus) resistant hybrids through intergeneric hybridization. Intergeneric hybridization was done involving nine Carica papaya cultivars as female and Vasconcellea cauliflora as male. To break the intergeneric hybridization barrier, various nutrient combinations were used. Among the combinations used, sucrose 5%, sucrose 5% + boron 0.5% and sucrose 5% + CaCl₂ 0.5% improved the fruit set and seed set percentage. A total number of 1197 flowers were pollinated and 308 fruits were obtained. On extraction, 721 seeds were obtained from CO 7, Pusa Nanha and CP 50. Out of 721 F₀ seeds (crossed seeds) sown, 419 seeds germinated and artificial screening for PRSV was carried out 27 days after sap inoculation. Out of 29 F₁ hybrid plants from CO 7 x V. cauliflora cross, only six plants namely CO 7V1 to CO 7V6 were found free from PRSV symptoms. Similarly, out of 55 F₁ hybrids from cross involving Pusa Nanha x V. cauliflora only 23 plants namely PNV1 to PNV23 were found free from the symptoms and 70 plants namely CPV1 to CPV70 out of 335 plants of CP50 x V. cauliflora cross were found free from PRSV symptoms. Among the crosses, Pusa Nanha x V. cauliflora had higher yield under PRSV infected conditions, however, total soluble solids and total sugars were found lesser than the CO 7 x V. cauliflora cross. The hybridity of the progenies were confirmed by using ISSR (Inter Simple Sequence Repeats) primers by the amplification of DNA from progenies and their parents. ISSR primers UBC 856, UBC807 and ISSR primer combinations UBC 856-817, UBC 810-817, UBC 861-817, UBC 856-810, UBC 861-810 and UBC 856-817 clearly amplified specific bands of the male parent, which were present in F₁ progenies, but it was absent in female parents.     

Identifying the markers and tagging a leucine-rich repeat receptor-like kinase gene for resistance to anthracnose disease in vegetable cowpea [Vigna unguiculata (L.) Walp.]

Anthracnose caused by the fungus Colletotrichum lindemuthianum is the most destructive disease of cowpea. Field-type cowpeas show various levels of resistance, whereas pole-type vegetable cowpeas are highly susceptible. Transfer of resistance available in field types to vegetable types is a major breeding objective in cowpea. This paper details the development of an F₂ mapping population by crossing field-type cowpea variety Kanakamony (Vigna unguiculata ssp. cylindrica) with pole-type vegetable cowpea variety Sharika (Vigna unguiculata ssp. sesquipedalis), screening this population with artificial inoculation and Bulked Segregant Analysis (BSA) with random marker systems Random Amplified Polymorphic DNA (RAPD) and Inter-Simple Sequence Repeats (ISSR); the objective is to identify the markers linked with major resistance-contributing genes. RAPD primer OPA02 has yielded marker at 850 bp in susceptible genome, whereas ISSR primers UBC810 and UBC811 have yielded markers at 1.4 kb and 1.5 kb respectively in resistant genomes. These markers were reproducible and their linkage with resistance and suitability in marker assisted selection (MAS) were confirmed through co-segregation analysis in F₃ population. UBC811 marker was eluted, cloned on pGEM-T, and sequenced. The sequence had shown that this marker is anchored on LRR receptor-like serine/t\hreonine protein kinase gene which could be involved in the resistance mechanism.     

与辣椒抗蚜虫基因相关的RAPD标记筛选及其SCAR转化

以辣椒抗蚜虫野生种质03-C79′与感蚜品种A0003杂交并自交得到F2为试材,应用RAPD分子标记技术和BSA混合群体分组分析法寻找与辣椒抗蚜虫基因相关的分子标记。从200条RAPD引物中筛选到1个辣椒抗蚜虫性状特异的多态性引物RA750。经F2单株验证,RA750的扩增片段在抗蚜虫植株中稳定出现,而在感蚜植株中没有。利用JoinmapV3.0软件计算标记与基因间的遗传距离为6.03 cM。根据对该片段的序列分析结果重新设计了1对引物,将RAPD标记转化成了SCAR标记,并命名为RA750-S。