刺槐Genomic-SSR与EST-SSR遗传差异性分析

[目的]对刺槐Genomic-SSR与EST-SSR的遗传差异性进行研究,为今后刺槐遗传多样性分析等育种相关研究中合理选用不同来源的SSR分子标记奠定基础。[方法]选取来自美国四个不同采集地的种子育出的12个刺槐个体,试剂盒提取DNA后分别利用9对Genomic-SSR引物和9对EST-SSR引物进行扩增,并采取毛细管电泳检测扩增产物。利用所得条带信息及相关软件对两种SSR分子标记进行多态性、遗传相似系数相关性以及聚类等方面的比较分析。[结果]刺槐Genomic-SSR平均检测到的条带数为6.0、Shannon多样性指数为1.3833、观测杂合度为0.5749、期望杂合度为0.6832；EST-SSR平均检测到的条带数为5.1、Shannon多样性指数为1.2711、观测杂合度为0.5648、期望杂合度为0.6526。由Genomic-SSR计算得到的个体间的遗传相似系数以及聚类结果与两种SSR标记综合计算得到的遗传相似系数和聚类结果更为相似。[结论]刺槐Genomic-SSR与EST-SSR存在一定的遗传差异性,但差异并不显著；刺槐Genomic-SSR能更加准确地揭示基因型之间的遗传关系；刺槐EST-SSR具有相对较强的保守性。     

马尾松二代育种亲本主要生长性状和ISSR遗传变异

本研究以国家马尾松种质资源库(浙江省淳安县姥山林场)马尾松二代核心育种群体内的36个亲本无性系为材料,研究其主要生长性状及ISSR遗传变异。马尾松二代育种亲本生长性状总体上优良,但生长性状在亲本无性系之间存在显著差异,表明二代育种群体具有丰富的表型性状变异。利用15个ISSR标记对36个二代亲本无性系进行分析,多态位点百分率(PPL)为83.33%。Nei's基因多样性(HE)为0.328,Shannon信息多样性指数(I)为0.476,亲本间分子遗传距离的范围是0.24～0.82,平均为0.414,在分子水平上证明马尾松二代育种群体遗传多样性较高,具有较好的高世代遗传改良潜力。基于Nei's无偏遗传距离进行UPGMA聚类,36个无性系中的35个聚在5个类群内,聚类结果与马尾松二代亲本的遗传背景、系谱关系及产地纬度部分吻合,证实了应用ISSR标记开展马尾松分子辅助育种的可行性。本研究为合理选配二代亲本、开展杂交育种提供一定借鉴。     

基于SSR标记的80份烟草种质指纹图谱的构建及遗传多样性分析

利用均匀分布于烟草24个连锁群上的48个SSR标记对80份烟草材料进行分析。结果显示,48个SSR标记共扩增出211个等位基因,平均每个标记4.396个,Shannon′s信息指数I为1.034,多态信息含量值为0.229~0.905,观测杂合度（H_o）、期望杂合度（H_e）和Nei′s多样性指数（H）平均值分别为0.320、0.572、0.431。聚类结果表明,在遗传距离为0.68时可以将80份烟草种质分为2个类群。5个烟草居群间的遗传一致度在0.643~0.765范围内,遗传距离分布在0.268~0.442。筛选4对核心引物构建了不同烟草种质资源的数字指纹图谱, 可将这80份烟草种质资源全部区分开。本研究在分子水平上为筛选优质烟草种质资源、挖掘重要基因以及拓宽烟草育种遗传基础等工作提供科学依据。     

利用新型分子标记EST-SSR鉴定湖北省内的主栽黑杨品种

EST-SSR标记是基于表达序列标签(expressed sequence tags,EST)数据信息,并结合SSR标记特点开发出的一种新型分子标记.本文利用12对EST.SSR标记对湖北省内目前主要栽培的8个黑杨品种进行分子鉴定,同时对不同EST.SSR位点进行相关遗传参数分析.品种鉴定结果表明,仅需4对EST-SSR引物即可将8个亲缘关系较近的黑杨品种完全区别开来.遗传多样性分析显示,多态位点百分比为58.3%,平均有效等位基因数为2.09个,平均Shannon信息指数0.61,平均杂合度为0.35.这些结果表明,杨树EST-SSR标记完全可以应用于品种鉴定及群体遗传多样性分析.     

基于SSR的四川花生遗传多样性分析

[目的]研究旨在了解四川花生资源的遗传多样性及表型性状与分子标记的关联分析,为花生分子育种及资源库构建提供理论依据。[方法]鉴定分析57份不同来源花生资源材料的4个农艺性状及4个品质性状,并利用SSR标记研究57份花生材料的遗传多样性,对SSR标记与表型性状进行聚类及关联分析。[结果]表型性状鉴定结果显示花生资源的8个性状中,6个变异较大、多样性较好。67对SSR引物中,有效引物39对,获得多态性条带53条,平均每个引物扩增1.36条,平均Nei"s 基因多样性0.2288、平均Shannon"s 指数0.3695,最远遗传距离为0.51。SSR分子标记聚类分析将57份资源聚为2大类群,GLM分析发现多个与蛋白质含量、株高、含糖量的关联标记。[结论]研究结果表明,四川不同区域间花生遗传多样性较丰富,品种类型单一,聚类及关联分析结果可为四川花生突破性新品种选育的亲本选择提供提供重要参考。     

黄瓜种质资源形态学标记和SRAP标记的遗传多样性分析

旨在探索黄瓜种质资源的遗传多样性,为今后黄瓜选育优良品种提供可靠依据。以48份黄瓜种质资源为材料,用DPS软件对各品种的13个表型性状进行聚类分析,同时利用SRAP分子标记对其进行遗传多样性研究。结果表明:根据果实性状等形态学标记聚类分析得出,在阈值75.0处,可以将48份黄瓜种质资源分成5大类群。利用UPGMA法对SRAP分子标记结果聚类分析发现,黄瓜种质间的遗传相似系数在0.52~0.93之间,在遗传相似系数为0.64处,可以将其分成5大类群,形态学标记和SRAP分子标记的聚类结果存在较大差异。由此可见,SRAP分子标记较形态学标记能更好地分析品种间亲缘关系的远近,对黄瓜遗传多样性研究更具指导意义。     

我国茶树无性系品种遗传多样性和亲缘关系的ISSR分析

利用ISSR分子标记分析了我国36个主要茶树无性系品种的遗传多样性和亲缘关系。结果表明20个ISSR引物在供试品种中共扩增出368条谱带,其中多态性条带占总条带的99.7%,引物的多态性信息指数（PIC）平均达0.90。供试品种的基因多样性（H）和Shannon信息指数（I）分别为0.23和0.38。茶区内茶树品种的遗传多样性低于总体水平,江南和华南茶区主栽无性系品种的多样性高于西南茶区。AMOVA分析表明区域因素引起的变异（占5.6%）远小于品种因素（占94.4%）。供试品种间的相似系数介于0.58~0.84,平均为0.69,显示出我国茶树主栽品种的遗传基础已相对比较狭窄。ISSR聚类分析表明,中国台湾品种金萱与大陆品种的遗传距离较远,形成单独的个类。35个大陆品种聚成一个大类群,其中除宜红早形成独立的个类外,其他品种又聚为3个亚类群。亲缘关系树状图在分子水平上显示了我国主要茶树无性系品种间的亲缘关系,为今后茶树育种亲本的选配提供了理论依据。     

82份玉米自交系遗传多样性分析

对种质资源进行遗传关系的研究和遗传多样性评价是玉米育种研究的重要内容,采用形态标记和SSR分子标记2种方法对82份玉米自交系进行了遗传多样性分析。结果表明：用15个表型性状计算它们的欧式遗传距离,其平均遗传距离为50.57,变异范围为9.05~146.23,说明供试自交系遗传多样性丰富,以遗传距离38.06为界,将供试自交系分为6类,其聚类结果与其系谱来源的吻合程度较差;利用筛选出来的63对扩增条带清晰、多态性明显的SSR引物,在供试自交系中共检测出等位基因变异601个,每对引物检测到4~24个等位基因,平均为9.5个,每个位点多态性信息量(PIC值)变幅为0.4546~0.9169,平均为0.7529,遗传相似系数为0.5441~0.9334,平均为0.6673,以遗传相似系数0.6735为界,将82份自交系分为七大类,属于五大常见类群的自交系占84.1%,其中Reid群占32．9%,PB群占23.2%,Lancaster群占13.4%,塘四平头群占7.3%,旅大红骨群占7.3%,其他2个类群分别占11．0%和4.9%,其聚类结果与其系谱来源的吻合程度较高。     

基于RAPD标记的芥蓝种质资源遗传多样性分析

调查了44份芥蓝种质的植物学性状,并利用RAPD分子标记分析了其遗传多样性。结果从150条随机引物中筛选出20个引物,20个引物共扩增出177条谱带,其中多态谱带105条,平均每个引物扩增出8.9条谱带和5.3条多态性谱,多态性比率为59.32%。基于RAPD标记,利用NTSYS-pc2.11构建了聚类树状图谱,遗传距离为0.70时,44份芥蓝资源可聚成六大类群。芥蓝种质存在着一定的遗传多样性,但原产华南而且主要产区也在华南,遗传多样性要小于芸薹属其他蔬菜。     

哲罗鱼生长速度相关性状的微卫星标记筛选

利用22个微卫星标记对随机交配的野生哲罗鱼F1176个个体的生长速度相关性状(体质量、全长、体高、体宽、头长、头宽和眼间距)进行分析,22个微卫星标记共检测到67个等位基因,各座位的等位基因2～6个,片段大小为109～452 bp,平均观测杂合度和期望杂合度分别为0.583 5和0.564 0,22个位点均有不同程度的偏离Hardy-Weiberg平衡。利用SPSS软件对7个性状正态分布检测发现,7个性状都显示出连续变异的特点,属典型的数量性状或者多基因遗传,符合正态分布;同时用GLM程序对22个微卫星标记与生长速度相关性状进行相关性分析,并将与性状显著相关的标记进行基因型之间的多重比较,结果显示,22个微卫星座位中,有11个标记与至少一种性状显著相关,其中与全长相关的标记最少为4个,与体高和体宽相关的标记最多的为7个;多重比较可知每个标记中同一性状均有部分基因型差异达到显著水平,不同标记所代表均值也有些差异。上述这些标记为进一步的分子标记辅助育种提供了基础。     

基于SSR、InDel分子标记的红甜菜遗传多样性分析

本研究旨在了解21份国内外红甜菜品种的遗传多样性及其演化地位。利用Indel和SSR分子标记，对21份红甜菜遗传多样性研究。结果表明，InDel标记共得到25个基因位点，24个位点具有多态性，Shannon’s信息指数平均值为0.583，Nei’s基因多样性指数平均值为0.370，等位基因K为2~4个，平均2.4个，平均观测杂合度为0.235，期望杂合度0.398，多态信息含量PIC值为0.330；SSR标记共扩增出28个基因位点，全部具有多态性，平均每个标记扩增出3.5个多态性基因位点，Shannon’s 信息指数平均值为0.926，Nei’s基因多样性指数平均值为0.548，等位基因K为2~7个，平均4.625个，平均观测杂合度0.399，期望杂合度0.587，多态信息含量PIC值为0.523。本研究发现红甜菜品种大多因亲缘关系较近，因此在选育新品种时，建议尽量选择遗传距离较远的品种作为亲本。     

不结球白菜形态性状及SSR遗传多样性关联分析

分析不结球白菜遗传多样性,筛选与形态性状相关联且具有多态性的标记位点,为不结球白菜的分子辅助育种选择、种质资源的利用和新品种选育提供依据。本研究以类型差异大、有南方特色的54份种质资源为材料,对其6个质量性状和5个数量性状进行3次田间调查,从40对引物中筛选出25对SSR引物进行遗传多样性分析,结合形态性状对54份不结球白菜种质资源进行统计分析。结果表明:6个质量性状间具有丰富的多样性,5个数量性状间有显著的差异性,其中SA变异系数最大,FC与L、W和SA呈显著正相关;25对SSR引物共检测到72个多态性位点,平均每对SSR引物产生2.88个多态性位点,PIC平均值为0.3831,说明不结球白菜SSR具有丰富的遗传多样性;54份不结球白菜的遗传相似系数变化范围为0.492~0.963,平均为0.742,在遗传距离0.6处可将供试材料划分为3个类群;通过关联分析发现5个与数量性状关联的标记位点,各标记的表型变异解释率在0.4657~0.6329,平均为0.5393。本试验获得的5个标记位点可为不结球白菜分子标记育种提供依据。     

基于形态学性状和SSR标记的花生品种遗传多样性分析和特异性鉴定

以101份南方花生区试品种为材料,利用形态学性状和SSR标记进行品种遗传多样性分析和特异性鉴定。结果表明, 29个形态学性状中有7个无多样性,其余22个的多样性指数为0.23～0.77,平均为0.43。在相似系数为0.76处,将供试品种划分为七大类群,同一育种单位的品种倾向于聚在一起。用40个SSR标记共检测出167个等位基因,单个标记检测的等位基因数2～6个,平均为4.18个。标记的多态性信息量(PIC)差异较大,最大为0.79,最小为0.26,平均为0.55。在相似系数为0.70处,供试品种可被划分为六大类群,同一省份育成的品种多聚为一类。Mantel检验发现品种间的形态学性状和SSR标记的相似系数矩阵相关性弱(r=0.36),SSR标记无法取代形态学性状单独用于花生品种特异性鉴定,但两者相结合能有效提高花生品种特异性鉴定的准确性。     

Molecular tools for clone identification: the case of the grapevine cultivar 'Traminer'

In viticulture, biotype identification problems have traditionally been solved using ampelography, ampelometry and chemical traits analysis. However, these tools have resulted in several false attributions, in particular when used at the clonal level. The availability of relatively cheap, reliable and reproducible tools to identify genetic differences at the clonal level would greatly facilitate the work of clonal patenting. In this work, 24 accessions of ‘Traminer’ cultivars were characterized using molecular markers. Three different approaches were applied: simple sequence repeats (SSR), amplified fragment length polymorphism (AFLP) and methyl‐sensitive amplified length polymorphism (MSAP). Results showed that SSRs were not a powerful tool for clonal distinction. In contrast, the AFLP technique was able to distinguish 16 out of the 24 cultivars, even though the average similarity was high (97.1%). The MSAP technique was used to evaluate qualitative differences in the degree of DNA methylation among clones. Results suggest that morphological differences among clones are probably due to the synergetic effect of genetic and epigenetic modifications, and that clonal identification could be greatly improved using molecular tools such as AFLP and MSAP.     

基于SSR标记的芋头种质资源遗传多样性分析及抗疫病鉴定

为了了解不同来源芋头种质资源的遗传多样性及其对疫病的抗性，本研究从100对SSR引物中筛选出21对多态丰富的SSR引物，并将其用于109份芋头资源的遗传多样性分析，同时采用人工接种芋头疫霉菌的方法对这些资源进性行了抗性评价。结果表明，21对SSR引物在109份芋头资源中共扩增出71个等位基因，变幅在2~4个，平均有效等位基因数3.38；观测杂合度(Ho)变化范围为0.0481~0.8900，平均为0.3483；预期杂合度(He)的变化范围为0.0841~0.6709，平均为0.3794；种群平均Shannon遗传多样性指数为0.6733；遗传距离在0~1之间，平均遗传距离为0.6014。在分支距离为0.49处，可将供试芋头资源分为3个类群。60%以上的抗性资源集中于I和II类群。本研究可为筛选、鉴定优良芋头种质资源及遗传育种工作提供科学依据。     

楸树EST-SSR标记开发及种质资源的遗传多样性分析

利用转录组数据开发SSR标记是一种经济高效的DNA分子标记开发策略。本研究利用高通量测序技术开发楸树(Catalpa bungei)EST-SSR标记,了解其在转录组序列中的分布类型,并利用41对多态性较好的引物对来自安徽(AH)、河南(HN)、湖北(HB)和山东(SD)的4个楸树居群的48个无性系的遗传多样性进行分析。结果表明:在大于1kb的14634条序列中,共鉴定出3999个SSR位点,580条序列包含2个及以上位点;单核苷酸重复(1957,48.94%)是最常见的SSR类型,其次是二核苷酸重复(1164,29.11%),三核苷酸重复(834,20.86%),四核苷酸重复(41,1.03%)和五核苷酸重复(3,0.08%);共扩增出243个等位基因,每个位点的等位基因数为3~13个,平均为4.85个,观测杂合度为0.14?1.00,平均为0.63,期望杂合度为0.42?0.84,平均为0.67,大部分位点偏离哈迪-温伯格平衡;AH和HN居群在有效等位基因数和期望杂合度的数值较高,表明其遗传多样性较丰富;分子方差分析(AMOVA)表明,遗传变异主要发生在居群内。本研究为楸树及同属其他树种种质资源鉴定、遗传多样性提供依据,且在指导楸树良种选育等方面具有重要的意义。     

Genetic diversity in red clover (Trifolium pratense L.) revealed by morphological and microsatellite (SSR) markers

The NPGS-USDA core collection with 85 accessions of red clover, an important forage species, is little described. The goal of the present study was to evaluate the diversity of a set of accessions from the core collection at the morphological and molecular level in order to extract some valuable accessions for Brazilian red clover breeding programs. Twenty-one morphological traits, collected in field and greenhouse in South Brazil, and seven SSR markers were used to describe 57 accessions from the U.S. core collection and one population cultivated in Southern Brazil. Variation between accessions was large for most of the 21 morphological traits. A cluster analysis based on the morphological traits revealed five distinct clusters that separated the populations according to flowering earliness, as already described, but also according to persistency, growth habit and dry matter productivity. Over seven SSR loci, the number of alleles averaged 11.1 alleles per locus. Genetic diversity measured with SSR markers was high, with a mean expected heterozygosity of 0.86. An analysis of molecular variance revealed that the largest proportion of variation (83.6%) resided at the within population level. Although the molecular markers also separated accessions into five clusters, there was no coincidence between the composition of groups found with morphological and molecular data. Use of genetic diversity in breeding programs requires to use the most promising populations, to combine positive traits such as persistency and forage yield, and probably to use within population variation to detect valuable genotypes that could be used as parents of synthetic varieties.     

Comparison of phenotypic and molecular marker-based classifications of hard red winter wheat cultivars

Genetic diversity is the basis for successful crop improvement and can be estimated by different methods. The objectives of this study were to estimate the genetic diversity of 30 ancestral to modern hard red winter wheat (Triticum aestivum L.) cultivars adapted to the Northern Great Plains using pedigree information, morphological traits (agronomic measurements from six environments), end-use quality traits (micro-quality assays on 50 g grain or milled flour samples for the six environments), and molecular markers (seed storage proteins separated using SDS-PAGE, 51 SSRs, and 23 SRAP DNA markers), and to determine the relationships of genetic distance estimates obtained from these methods. Relationships among diversity estimates were determined using simple (Pearson) and rank (Spearman) correlation coefficients between distance estimates and by clustering cultivars using genetic-distances for different traits. All methods found a wide range in genetic diversity. The genetic distance estimates based on pedigree had the highest values due to possible over-estimation arising from model assumptions. The genetic diversity estimates based on seed storage protein were lowest because they were the major determinants of end-use quality, which is a highly selected trait. In general, the diversity estimates from each of the methods were positively correlated at a low level with the exceptions of SRAP diversity estimates being independent of morphologic traits (simple correlation), SDS-PAGE, and SSR diversity estimates (rank correlation). However, SSR markers, thought to be among the most efficient markers for estimating genetic diversity, were most highly correlated with seed storage proteins. The procedures used to accurately estimate genetic diversity will depend largely upon the tools available to the researcher and their application to the breeding scheme.     

Genetic diversity assessment and genotype identification in sugarcane based on DNA markers and morphological traits

Sugarcane is known for its highly complex genetics and more knowledge is needed for better use and conservation of genetic materials. In order to identify genotypes and to assess genetic diversity, diverse data sets such as morphological and molecular markers are used as a general approach. To evaluate the usefulness of different markers, important sugarcane genotypes in Argentina were characterized by AFLP, SSR and morphological traits. All genotypes characterized were grouped in one main cluster in dendrograms using two independent softwares. Interestingly, local genotypes grouped together with USA varieties and no clear genetic differentiation could be found probably due to intensive germplasm exchange between these breeding programs. The molecular markers tested were useful for genetic diversity assessment as well as for genotype identification. These markers should be included in the internationally established characters for sugarcane variety protection as they give a better view on whole genome complexity. Additionally, genetic similarities obtained from molecular markers will provide more accurate information to breeders than the pedigree method, especially when considering the asymmetric genetic inheritance of sugarcane. Morphological traits are valuable tools to identify genotypes since they reflect external resemblance more than genetic relatedness. When they were combined with molecular markers the dendogram obtained revealed genetic relationships and the genetic diversity was better estimated. In summary, both methods appear to be useful, complementing each other and should be used together to assist sugarcane breeders in estimating genetic diversity, electing parents for crossings, identifying superior lines and to protect intellectual property rights.     

马氏珠母贝选系F4遗传结构和亲缘关系建分析

为了有效避免因近交引起性状退化,本实验利用微卫星标记分析了选系F4的遗传结构,并估计了其有效亲本的数量。2010年9月,从快速生长系F3选择亲本进行子代繁殖,雌雄亲本数量分别为42和38个,人工解剖授精,按照常规技术进行幼体培育和海区养殖。2011年6月,从选系F4随机选取90个个体,利用45对微卫星引物进行遗传结构分析和有效亲本数量的推断。结果表明：45个微卫星位点共检测到186个等位基因,每个位点的等位基因数在2~9之间,平均每个位点有4.13个等位基因。平均观察杂合度为0.543;平均期望杂合度为0.542;平均Shannon多样性指数为1.012;平均PIC值为0.491。根据基因频率的似然率算法成功的推断出该群体含有30个全同胞家系,用于繁殖选系F4有效亲本数量为60个,有75%的亲本参与了繁殖。世代之间近交系数增量△F=0.83%,F4代群体的近交系数F=3.27%。本研究结果表明：（1）该选系F4具有较高的遗传变异;（2）利用微卫星标记能有效推算选系的有效亲本数量,同时为构建继代群体提供技术支持。