青藏高原早熟甘蓝型春油菜遗传资源创新研究

摘要：利用SSR和SRAP两种分子标记研究了71份实验材料的遗传差异及其亲缘关系。结果表明：(1) 29对SSR标记共扩增出121条清晰地带,多态性位点分别占总扩增位点的97.5%,而27对SRAP引物扩增出175条清晰地带,多态性比率为70.3%。（2）两种标记混合聚类结果表明,在相似系数0.566处所有材料可以分为A、B两个大类群。B类在相似系数0.620处又可分为六个亚类,10个天然双低早熟甘蓝型品系、两个甘蓝型亲本和四个新型品系聚在第Ⅰ亚类中,其余的51个新型甘蓝型油菜品系分别聚在其他五个亚类中。（3）对55份新型品系进行遗传成分分析,结果表明,每个品系都含有四种带型,不同品系中不同带型所占总带数的比率不同;对新型甘蓝型品系中含有白菜型亲本特有的带所占比率分别与其和对应的甘蓝型和白菜型亲本之间的遗传距离进行相关分析,结果表明新型甘蓝型品系中白菜型亲本特有的带所占比率与其和白菜型亲本之间的遗传距离为负相关（-0.52）,达到极显著水平;与其和甘蓝型亲本间的遗传距离为正相关（0.31）,达到显著水平。（4）对实验材料之间的遗传距离及其来源进行分析（除与两个白菜型亲本间）,遗传距离排前20位的都来自新型品系之间或天然品系与新型之间,且最大的达到0.544。     

西藏牦牛SRAP遗传多样性及分类进化研究

用4对SRAP分子标记引物对西藏11个牦牛类群和四川麦洼牦牛的DNA进行扩增,研究其遗传多样性和分类关系。结果表明,在335头牦牛中,共得到29个基因位点,其中有19个多态位点,多态率占65.52%。12个牦牛群体间的Nei’s遗传多样性和Shannon多样性指数分别为0.048 2和0.073 4,遗传相似系数在0.781 1-0.989 1。巴青牦牛和康布牦牛的遗传多样性指数较其他类群高,分别为0.095 5和0.090 0;桑日牦牛类群的遗传多样性指数最低,仅为0.008 5。这些结果表明,12个牦牛类群的SRAP遗传多样性较低。根据Nei’s遗传距离,利用UPGMAM构建聚类关系图结果显示,嘉黎牦牛、帕里牦牛、类乌齐牦牛、桑桑牦牛、康布牦牛、巴青牦牛、丁青牦牛、斯布牦牛和麦洼牦牛聚为一大类,然后依次才与桑日牦牛、工布江达牦牛和江达牦牛相聚在一起,显示在SRAP分子遗传标记所反映的牦牛基因组的遗传结构中,江达牦牛、工布江达牦牛和桑日牦牛与其他牦牛群体间的亲缘关系较远,牦牛的这种亲缘关系与其地理分布也不一致,说明这12个牦牛的起源、演化关系较复杂,有待于进一步研究分析。     

29个香蕉基因型遗传多样性的SRAP分析

采用SRAP分子标记技术对29个香蕉品种(系)的多样性进行研究,结果显示,64对SRAP引物中筛选出25个多态性较高的引物组合,共扩增出324条条带;UPGAM聚类图显示所有供试的29个香蕉品种(系)可分为2个类群且与基因型相一致;实验结果与形态、农艺性状标记分类基本一致。研究表明,SRAP技术可有效运用于香蕉基因型的遗传和育种研究。     

中国半夏属植物亲缘关系的SRAP分析

利用序列相关扩增多态性(SRAP)标记对中国半夏属植物5个种的亲缘关系进行了研究.38个引物组合在半夏属植物的5个种中共扩增出752条清晰的谱带,其中628条谱带具有多态性,多态性比率为83.51％,显示出较高的多态性比率;各物种间的遗传相似系数在0.6513～0.7312之间,聚类分析和主坐标分析结果表明,5种半夏属植物被聚为两大类:掌叶半夏单独聚为一类(Ⅱ),而其它4个种聚为一类(Ⅰ).第Ⅰ类可再分为A和B两个亚类:A亚类包括半夏和石蜘蛛;B亚类包括盾叶半夏和滴水珠.滴水珠和盾叶半夏的亲缘关系最近,其次是半夏和石蜘蛛,而掌叶半夏和其它4个种的亲缘关系都较远,这说明掌叶半夏与半夏属其它种呈姐妹群关系.本研究结果对我国半夏属植物资源的开发利用与保护具有重要意义.     

Genetic diversity and phylogenetic relationships among five endemic Pinus taxa (Pinaceae) of China as revealed by SRAP markers

The genetic diversity and phylogenetic relationships among five endemic Pinus taxa of China (Pinus tabulaeformis, P. tabulaeformis var. mukdensis, P. tabulaeformis f. shekanensis, Pinus massoniana and Pinus henryi) were studied by SRAP markers. Using 10 SRAP primer pairs, 247 bands were generated. The percent of polymorphic bands (94.8%), Nei's genetic diversity (0.2134), and Shannon's information index (0.3426) revealed a high level of genetic diversity at the genus-level. At the taxon level, P. tabulaeformis f. shekanensis and P. henryi showed a higher genetic diversity than the others. The coefficient of genetic differentiation among taxa (0.3332) indicated a higher level of genetic diversity within taxon, rather than among taxa. An estimate of gene flow among taxa was 1.0004 and implied a certain amount of gene exchange among taxa. The results of neighbor-joining cluster analysis and principal co-ordinate analysis revealed that P. tabulaeformis, P. tabulaeformis var. mukdensis and P. tabulaeformis f. shekanensis were conspecific, which was in agreement with the traditional classification. Phylogenetic relationships analysis also indicated that P. henryi might be a distinct species closely related to P. tabulaeformis.     

RAPD及SRAP两种分子标记技术对苦瓜杂交种纯度的鉴定分析

以F1代苦瓜杂交种如玉11号及其亲本为材料,利用RAPD及SRAP两种分子标记技术对这3种苦瓜基因组DNA进行比较分析,以获得该杂交种及其亲本（或母本）差异目的基因片段。经过多次对该3种苦瓜叶片DNA提取,PCR扩增及其PCR产物的琼脂糖凝胶电泳分析,在供试的46个RAPD引物及121对SRAP引物中,筛选出1个RAPD引物及1对SRAP引物能区分该苦瓜杂交种及其母本种子,通过进一步验证分析,证明该两种分子标记的特异引物可作为如玉11号苦瓜杂交种子的纯度鉴定之用。     

基于SRAP标记的薏苡种质资源遗传多样性及DNA指纹图谱构建

利用SRAP分子标记对从各主要产地收集到的90份薏苡种质进行遗传多样性分析,其中68份收集于福建省,6份来自中国台湾,16份来自浙江、辽宁、山东、河南、云南、江苏、湖南、广东、上海等省(市)。结果表明,从88对SRAP引物组合中筛选出26对引物进行SRAP扩增,共扩增出185条带,其中具有多态性的有157,占总数的84.86%,表明90份薏苡种质表现出丰富的遗传多样性。基于SRAP标记利用系统聚类法将90份薏苡种质资源分为4大类,与形态性状分类结果有一定的相似性;利用16对SRAP引物构建了73份薏苡种质资源的DNA指纹图谱,为薏苡遗传研究、品种选育与资源保护提供了依据。     

茄子种质遗传多样性及群体结构的SRAP分析

采用SRAP标记法对183份茄子(Solanum melongena)种质资源的遗传关系和群体结构进行分析。结果表明, 33对多态性SRAP引物组合共扩增出215条清晰稳定的多态性条带, 平均每对引物组合产生7条多态性条带。183份茄子种质的遗传相似系数介于0.276-0.813之间, 平均值为0.623, 表明茄子种质资源间遗传背景存在一定的差异。在遗传距离为0.345 6处可将183份茄子种质分为4组。通过群体结构分析可将种质划分为4个群体, 不同群体间的界限十分明显, 且群体间的基因渗透较高。     

Map-based cloning of genes encoding key enzymes for pigment synthesis in Auricularia cornea

Color is an important quality attribute of fungi, and a useful marker for classification, genetic, and molecular research. However, there is much debate over which enzymes play key regulatory roles in pigment synthesis pathways among different fungi and even within the same species. Auricularia cornea is the most widely cultivated mushroom in the genus Auricularia; 1.834 million tons of this mushroom were produced in 2016 in China. Thus, systematic studies on its color inheritance and the genes encoding key enzymes for pigment synthesis have high scientific and economic value. In this study, the white strain ACW001 and the purple strain ACP004 of A. cornea were used as dikaryotic parents. Selfing populations of ACW001 and ACP004 were constructed with their monokaryotic strains. The fruiting body color of the two populations was consistent with that of their parents, confirming that the two parents were color homozygotes. All strains in the hybrid population of the two parents produced purple fruiting bodies. A robust hybrid strain (ACW001-33×ACP004-33) was selected from the hybrid population, and 87 monokaryotic strains of ACW001-33×ACP004-33 were obtained as a mapping population. Finally, a testcross population was constructed by crossing the mapping population with the test strain ACW001-9. The color genotype of each monokaryotic strain in the mapping population was identified by a fruiting test. The genomes of the two monokaryotic strains ACW001-33 and ACP004-33 were sequenced, and then simple sequence repeat (SSR) and sequence-related amplified polymorphism (SRAP) molecular marker primers were developed. Then, 88 pairs of primers that could distinguish the genotypes of the mapping population were used to construct a genetic linkage map. The genetic linkage map consisted of 12 linkage groups (LGs) spanning 1315.2 cM. The color control locus was preliminarily located at 24.5 cM of the 11th LG. Fine-mapping primers were designed based on sequence differences between ACW001-33 and ACP004-33 in the primary location region. Four color control candidate genes were located in an 8.2-kb region of ACW001-33_contig733 and a 9.2-kb region of ACP004-33_contig802. Homologous alignment and prediction of conserved domain analyses indicated that two of the color control candidate genes encoded proteins with unknown function, and the other two, ACP004_g11815 and ACP004_g11816, encoded glutamyl aminotransferases. These two genes were consecutively arranged on ACP004-33_contig802, and were likely to encode key enzymes in the γ-glutamine-4-hydroxy-benzoate (GHB) pigment synthesis pathway. Primers were designed from the flanking sequences of the two genes and used to analyze the testcross population. Products were amplified only from the 30 testcross strains with purple fruiting bodies, confirming the accuracy of the localization results. We discuss the deficiencies and advantages of map-based cloning in fungi vs. plants, and summarize the steps and requirements of the map-based cloning method for fungi. This study has provided novel ideas and methods for locating functional genes in fungi.