可可西里自然保护区藏羚羊的微卫星多态性研究

藏羚羊是我国特有的珍稀濒危动物,对其开展遗传多样性的研究具有非常重要的科学价值。为了获取足够的遗传信息并进一步研究藏羚羊在核基因水平上的遗传多样性,对来自可可西里地区的75个藏羚羊干皮张样本进行了微卫星遗传多样性研究。研究从来自牛和绵羊的25个微卫星基因座中筛选到9个具有高度多态性的微卫星基因座(MCM38,MNS64,IOBT395,MCMAI,TGLA68,BM1329,BMS1341,BM3501和MB066)。用非变性聚丙烯凝胶电泳检测微卫星的PCR扩增产物,计算了这9个微卫星基因座的等位基因频率、多态信息含量、基因杂合度等指标并估算了种群数量。结果在75只藏羚羊中共检测到85个等位基因,9个微卫星基因座的等位基因数为7~12个,平均每个基因座检测到9.4个等位基因,有效等位基因数为处于4.676~9.169之间,平均为6.519;基因频率分布在0.007~0.313之间,多态信息含量在0.753~0.881之间,平均为0.818;观察杂合度为0.791~0.897,平均为0.844,期望杂合度为0.786~0.891之间,平均为0.838±0.0132,各基因座观察杂合度与期望杂合度比较接近。固定指数为-0.269~-0.097,平均为-0.163。Shannon’s指数为1.660~2.315,平均为1.990。种群数量的估算结果显示这75个体均来自同一种群。结果表明该种群在核基因水平仍具有丰富的遗传多样性。     

长江中上游两个鲢群体遗传变异的微卫星分析

对长江中上游2个鲢群体使用39个微卫星标记进行了遗传多样性分析, 计算并统计了平均观测等位基因数、平均有效等位基因数、多态信息含量、遗传杂合度、Hardy-Weinberg平衡偏离指数、遗传相似系数、遗传距离等遗传参数。结果表明: 万州鲢和监利鲢群体所检测微卫星位点的平均观测等位基因数分别为6.128和4.974; 平均有效等位基因数分别为4.107和3.395; 多态位点百分率分别为100和94.87; 39个微卫星标记共有等位基因259个, 173个等位基因为两群体所共有; 多态微卫星位点的PIC在0.077~0.865之间变动,平均为0.617; 两群体所检测位点平均观测杂合度为0.834和0.775, 平均期望杂合度为0.713和0.623; 两个群体间的遗传相似系数为0.618, 群体间的遗传距离为0.482。结果显示长江中上游两个鲢群体间存在显著遗传分化, 应隶属于不同的种群。     

中国圈养林麝微卫星DNA多样性研究

本文利用13对微卫星标记,对我国3个核心种源地（巴山、秦岭、川西高原）圈养林麝种群进行遗传多样性和遗传结构分析。在167份样品中共检测到142个等位基因（Na）,每个位点等位基因数介于7~16,均值为10.92,平均有效等位基因数（Ne）为6.3730,期望杂合度（He）和观测杂合度（Ho）均值分别为0.8302和0.3897。这些圈养林麝种群遗传多样性水平较高,但较低的观测杂合度表明圈养群体存在近交现象。两两群体间的Fst 值和AMOVA分析结果均表明种群之间分化程度不明显。群体遗传结构分析显示,全部样本聚为3个遗传簇（最佳K值=3）,其主体与3个地理来源相符,但种群间存在基因渗透现象。本研究中的秦岭种群遗传变异最为丰富,可以作为种质改良的基因池。     

草鱼种群SSR分析中样本量及标记数量对遗传多度的影响

利用45对微卫星分子标记（SSR）,以草鱼(Ctenopharyngodon idellus)自然群体为实验材料,探讨野生群体遗传多样性研究中所需的最适样本量与标记量。实验设置6个样本量梯度,9个标记量梯度。对等位基因数（Na）、有效等位基因数（Ne）、观察杂合度（Ho）、期望杂合度（He）等遗传多样性指标的变化趋势进行统计分析。结果表明,样本量、微卫星标记的数量和多态性水平对群体遗传多样性均有较大的影响,其中等位基因数与样本量大小呈显著正相关,而杂合度随标记量的增多而剧烈波动。当取样量大于40,标记量大于25时,各遗传参数值趋于稳定。因此,在应用微卫星标记对水产动物自然群体的遗传学研究中,要根据所研究种类的特点,尽可能采样40尾以上,采用25个以上标记,避免由人为选择的偏差对群体遗传多样性水平的正确评估所造成的影响。同时根据上述研究结果,对陕西草鱼自然群体进行了遗传多样性的评估,结果显示该群体平均等位基因数（MNA）、平均有效等位基因数、平均观测杂合度、平均期望杂合度分别为7.26、4.21、0.73、0.68,认为该群体具有较高的遗传多样性。     

黑龙江省张广才岭北部小飞鼠的遗传多样性与种群历史动态

小飞鼠 (Pteromys volans) 为树栖夜行滑行类啮齿动物,在森林种子传播和维持生态系统平衡等方面发挥着重要的生态学作用。本研究利用mtDNA Cytb、控制区和nDNA微卫星3种分子标记,对黑龙江省张广才岭北部的小飞鼠种群进行遗传多样性与历史动态分析。检测出Cytb全序列 (1 140 bp) 的平均单倍型多样性为0.909,平均核苷酸多样性为0.616%;控制区全序列 (1 066 bp) 的平均单倍型多样性为0.945,平均核苷酸多样性为1.698%;微卫星检测出种群平均等位基因数13.167个,观测杂合度0.727,期望杂合度0.864,近交系数0.159。结果表明,小飞鼠种群遗传多样性丰富,但存在一定程度的杂合度不足和近亲繁殖;未检测到种群近期遗传瓶颈效应,种群内无遗传分化。高比例的稀有单倍型 (≥ 60%) 、低频率等位基因与近亲繁殖,提示未来种群面临遗传多样性下降的风险,建议加大对该物种的关注和保护力度。基于Cyt b基因的系统进化关系结果表明,小飞鼠存在3个明显的遗传谱系：远东、欧亚大陆北部和日本北海道,本研究中张广才岭和大兴安岭的样本单倍型归属为远东谱系。     

利用微卫星研究普洱亚洲象种群的遗传多样性

本文采用非损伤性取样法获得普洱地区亚洲象粪便样品113份,试剂盒法提取粪便基因组总DNA,利用9对微卫星引物对DNA进行特异性扩增,CERVUS 3.0软件基因分型得到49个独特的基因型。利用GenAlEx v6.5与Arlequin v3.5进行位点遗传信息检测和种群遗传多样性分析,结果表明:研究采用的所有微卫星位点均未偏离哈迪温伯格平衡,位点的平均等位基因数为3.111,平均香农信息指数为0.804,平均期望杂合度为0.468,平均观测杂合度为0.476。比较相同位点上不同地区亚洲象的遗传杂合度,表明普洱地区亚洲象的种群遗传多样性较低。     

基于粪便DNA的东北马鹿遗传多样性

东北马鹿(Cervus canadensis xanthopygus)为国家二级重点保护野生动物,近些年其种群数量急剧下降、分布区不断退缩、种群基因交流受阻,很多地区更是难觅其踪迹。亟需对其种群的遗传变化,特别是遗传多样性和近交衰退等种群遗传信息开展进一步评价,增强保护与管理的针对性。本研究在大、小兴安岭和长白山脉的6个重点研究区域,共收集409份疑似马鹿粪便样本。首先基于mtDNA Cyt b基因测序技术进行物种鉴定,并对鉴定为马鹿的阳性样本利用微卫星技术进行个体识别。结果共识别出172只东北马鹿个体;Cyt b基因序列共检测出14个变异位点和11个单倍型,单倍型多样性为0.849 (0.105～0.732),核苷酸多样性为0.678%(0.099%～0.775%)。10个微卫星位点检测出种群平均等位基因数为5.7 (5.2～7.2),有效等位基因数为3.3 (2.5～4.1),观测杂合度为0.687 (0.644～0.725),期望杂合度为0.619 (0.564～0.689),近交系数为-0.113 (-0.160～-0.037)。结果表明,东北马鹿种群遗传多样性处于中等水平,其...     

三个野生群体日本囊对虾遗传多样性的SSR分析

为了解野生种群日本囊对虾遗传分化和改良遗传育种,用SSR技术对福建厦门(XM)、广东湛江(ZJ)、广西北海(BH)3个地区野生日本囊对虾进行遗传多样性的研究。采用了10对微卫星引物对3个野生种群进行分析,10个微卫星位点在3个种群中均表现为高度的多态性,每个位点平均检测到3.87个等位基因;平均多态信息含量为0.5893;3个群体的观测杂合度分别为0.6243、0.5704、0.4661,全部群体观测杂合度平均为0.5536;期望杂合度分别为0.7193、0.6189、0.6226,全部群体平均期望杂合度为0.6536。这说明3个野生种群在10个微卫星位点上均具有丰富的遗传多样性。基于Nei's遗传距离的聚类分析显示厦门群体和湛江群体的遗传距离较近。     

近缘种扩增法对黑叶猴微卫星位点的筛选及特征分析(英文)

微卫星位点近缘种筛选法使得在探讨各种灵长类种群遗传结构和生殖策略上更加便捷。我们利用138条人类微卫星引物在黑叶猴中进行筛选,得到了23个具有多态性位点。在28个检测个体中,每个位点的等位基因数为3到9个,期望杂合度为0.62,观测杂合度为0.50,其中有7个位点偏离Hardy-Weinberg平衡,9个位点存在无效等位基因现象。但是各位点之间均未检测到连锁不平衡现象。这些位点将在黑叶猴种群遗传结构的研究中发挥重要作用。     

白颈长尾雉微卫星多态性的遗传学分析

采用7个微卫星位点遗传标记,对白颈长尾雉(Syrnaticus ellioti)4个地理种群105个个体进行了种群遗传分析。研究发现4个地理种群均偏离Hardy-Weinberg平衡,共检测到62个等位基因,平均等位基因数目为8.86;大多数微卫星位点的观察杂合度值较低,平均为0.504,要明显低于期望杂合度。7个位点的多态信息含量为0.549～0.860,平均为0.712。用无限等位基因模型、逐步突变模型和双相突变模型对4个地理种群的种群瓶颈效应检测,发现各种群近期内都经历过瓶颈效应的影响。地理种群之间的Fst值表明,贵州与湖南地理种群间的分化达到了极显著水平(P<0.001);由Nei氏的无偏遗传距离所构建的邻接树显示,贵州地理种群与湖南地理种群的遗传关系较远。微卫星对不同地理种群的分层分子变异分析(贵州地理种群对其他地理种群)发现:来自地理种群间和组群间的遗传变异量相对较小;而同一地理种群内个体之间的变异量较大(92.84%),且达到显著水平。     

用微卫星标记分析皱纹盘鲍群体的遗传变异

利用微卫星标记技术, 对皱纹盘鲍山东长岛和辽宁獐子岛的两个野生群体以及山东崆峒岛一个养殖群体的遗传变异进行了分析。对6个微卫星基因座的多态性进行了评估, 各基因座的多态信息含量(PIC)值均大于0.5, 适合对鲍群体遗传结构的分析。结果表明, 这6个基因座在3个皱纹盘鲍群体中共获得57个等位基因, 等位基因数(A)平均为9.50, 有效等位基因数(N_e)平均为5.8572, 平均杂合度观测值(H_o)和期望值(^H_e)分别为0.6925和0.7966; 两个野生群体的杂合度观测值(H_o)和期望值(H_e)均高于养殖群体。上述结果为保护和利用皱纹盘鲍的遗传多样性提供了依据。     

Importance of a pilot study for non-invasive genetic sampling: genotyping errors and population size estimation in red deer

Population size information is critical for managing endangered or harvested populations. Population size can now be estimated from non-invasive genetic sampling. However, pitfalls remain such as genotyping errors (allele dropout and false alleles at microsatellite loci). To evaluate the feasibility of non-invasive sampling (e.g., for population size estimation), a pilot study is required. Here, we present a pilot study consisting of (i) a genetic step to test loci amplification and to estimate allele frequencies and genotyping error rates when using faecal DNA, and (ii) a simulation step to quantify and minimise the effects of errors on estimates of population size. The pilot study was conducted on a population of red deer in a fenced natural area of 5440 ha, in France. Twelve microsatellite loci were tested for amplification and genotyping errors. The genotyping error rates for microsatellite loci were 0–0.83 (mean=0.2) for allele dropout rates and 0–0.14 (mean=0.02) for false allele rates, comparable to rates encountered in other non-invasive studies. Simulation results suggest we must conduct 6 PCR amplifications per sample (per locus) to achieve approximately 97% correct genotypes. The 3% error rate appears to have little influence on the accuracy and precision of population size estimation. This paper illustrates the importance of conducting a pilot study (including genotyping and simulations) when using non-invasive sampling to study threatened or managed populations.     

利用17个微卫星标记分析鳙鱼的遗传多样性

选用本实验室克隆的17个鳙鱼微卫星分子标记分析四川泸州和江西鄱阳湖的两个种群鳙鱼的遗传多样性及种质特性,计算和统计了杂合度、多态信息含量（PIC）、有效等位基因数、等位基因频率、遗传距离、遗传相似系数、Hardy-Weinberg平衡偏离指数等方面内容。结果表明：选择使用17个微卫星标记,其中有4个为单态标记,13个为多态标记。江西和四川鳙鱼群体每个微卫星位点的平均等位基因数分别为3.325及3.882,平均有效等位基因数分别为3.531及2.676,多态位点百分率分别为82.4及70.5, 17个微卫星标记共有等位基因71个,多态微卫星位点的PIC在0.114~0.960之间变动,平均为0.417 ,两群体位点平均观测杂合度为0.385和0.452,平均期望杂合度为0.360和0.422,两个群体间的遗传相似系数为0.897,群体间的遗传距离为0.109。     

镜鲤繁殖群体的遗传结构及微卫星标记与经济性状的相关性分析

选用35个多态性微卫星分子标记对天津换新良种场镜鲤一个繁殖群体的有效等位基因数(Ae)、观测杂合度(Ho)、期望杂合度(He)、多态信息含量(PIC) 等进行了检测, 以卡方检验估计群体Hardy-Weinberg平衡。结果表明：在35个基因座共检测到118个等位基因, 平均等位基因数为3.37个, 每个座位检测到的等位基因数2~7个不等, 平均有效等位基因数为2.16, 观测杂合度平均值0.431, 无偏期望杂合度的平均值为0.4736, 平均多态信息含量0.42, 说明这个群体属于中度多态, 遗传多样性水平不高。卡方检验的P值显示多于半数的位点都发生了偏离。并将35个基因座的不同基因型与个体的体重、体长值进行了连锁分析, 得到了4个与体重、体长连锁的基因型, 并将所得结果与鲤鱼体长性状QTL定位结果进行对比, 其中HLJ319标记与QTL定位结果基本一致。分析了几个严重偏离平衡的基因型, 并讨论出现这种现象的可能原因。     

Microsatellite analysis of genetic diversity in the Chinese alligator (<Emphasis Type="Italic">Alligator sinensis</Emphasis>) Changxing captive population

Chinese alligator (Alligator sinensis) is a critically endangered species endemic to China. In this study, the extent of genetic variation in the captive alligators of the Changxing Reserve Center was investigated using microsatellite markers derived from American alligators. Out of 22 loci employed, 21 were successfully amplified in the Chinese alligator. Sequence analysis showed loci in American alligators had a bigger average size than that of the Chinese alligators and the longest allele of an individual locus almost always existed in the species with longer stretch of repeat units. Eight of the 22 loci were found to be polymorphic with a total of 26 alleles present among 32 animals scored, yielding an average of 3.25 alleles per polymorphic locus. The expected heterozygosity (H _E) ranged at a moderate level from 0.4385 to 0.7163 in this population. Compared to that in the American alligators, a lower level of microsatellite diversity existed in the Changxing population as revealed by about 46% fewer alleles per locus and smaller H _E at the homologous loci. The average exclusion power and the ability to detect shared genotypes and multiple paternity were evaluated for those markers. Results suggested that when the polymorphic loci were combined, they could be sensitive markers in genetic diversity study and relatedness inference within the Chinese alligator populations. The level of genetic diversity present in the current Changxing population indicated an important resource to complement reintroductions based on the individuals from the other population. In addition, the microsatellite markers and their associated diversity characterized in this population could be utilized to further investigate the genetic status of this species.     

Assessment of population genetic structure in common wild rice Oryza rufipogon Griff. using microsatellite and allozyme markers

The genetic structure of five natural populations of common wild rice Oryza rufipogon Griff. from China, was investigated with 21 microsatellite loci and compared to estimates of genetic diversity and genetic differentiation detected by 22 allozyme loci. Microsatellite loci, as expected, have much higher levels of genetic diversity (mean values of A = 3.1, P = 73.3%, Ho = 0.358 and He = 0.345) than allozyme loci (mean values of A = 1.2, P = 12.7%, Ho = 0.020 and He = 0.030). Genetic differentiation detected by microsatellite loci ( FST = 0.468, mean I = 0.472) was higher than that for allozyme loci ( FST =0.388, mean I = 0.976). However, microsatellite markers showed less deviation from Hardy-Weinberg expectation (Wright's inbreeding coefficient FIS = -0.069) than do allozymes ( FIS = 0.337). These results suggest that microsatellite markers are powerful high-resolution tools for the accurate assessment of important parameters in population biology and conservation genetics of O. rufipogon, and offer advantages over allozyme markers.     

Sampling for Microsatellite-Based Population Genetic Studies: 25 to 30 Individuals per Population Is Enough to Accurately Estimate Allele Frequencies

One of the most common questions asked before starting a new population genetic study using microsatellite allele frequencies is “how many individuals do I need to sample from each population?” This question has previously been answered by addressing how many individuals are needed to detect all of the alleles present in a population (i.e. rarefaction based analyses). However, we argue that obtaining accurate allele frequencies and accurate estimates of diversity are much more important than detecting all of the alleles, given that very rare alleles (i.e. new mutations) are not very informative for assessing genetic diversity within a population or genetic structure among populations. Here we present a comparison of allele frequencies, expected heterozygosities and genetic distances between real and simulated populations by randomly subsampling 5–100 individuals from four empirical microsatellite genotype datasets (Formica lugubris, Sciurus vulgaris, Thalassarche melanophris, and Himantopus novaezelandia) to create 100 replicate datasets at each sample size. Despite differences in taxon (two birds, one mammal, one insect), population size, number of loci and polymorphism across loci, the degree of differences between simulated and empirical dataset allele frequencies, expected heterozygosities and pairwise F_ST values were almost identical among the four datasets at each sample size. Variability in allele frequency and expected heterozygosity among replicates decreased with increasing sample size, but these decreases were minimal above sample sizes of 25 to 30. Therefore, there appears to be little benefit in sampling more than 25 to 30 individuals per population for population genetic studies based on microsatellite allele frequencies.     

微卫星DNA标记探讨镜鲤的种群结构与遗传变异

采用30个微卫星分子标记, 对5个镜鲤群体的观测杂合度(^H_o)、期望杂合度(^H_e)、多态信息含量(PIC)和有效等位基因数(^A_e)等进行了遗传检测, 根据基因频率计算遗传相似系数和Nei氏标准遗传距离, 以c2检验估计Hardy-Weinberg平衡, 以近交系数(FST)和基因流(Nm)分析群体的遗传分化。同时, 使用PHYLIP3.63软件绘制基于Nei氏标准遗传距离的UPGMA聚类图, 并进行bootstrap自举检验验证进化树的可靠性。在德国镜鲤选育系(Scattered Cyprinus carpio L.)和来自4个不同养殖场(松浦、东岗、奉城和辽中)的德国镜鲤群体中共检测到7 083个扩增片段, 长度在102 ~ 446 bp之间, 在群体内扩增出等位基因1~16个不等, 共计356个等位基因。结果表明: (1)5个群体检测的有效等位基因数在1.07~12.30个不等, 平均多态信息含量为0.74、0.74、0.69、0.75和0.75, 无偏期望杂合度的平均值为0.74、0.78、0.70、0.76和0.78, 说明这几个群体属于高度多态, 遗传多样性水平较高。(2)群体间相似系数在0.52以上, 相似性较高。聚类分析显示, 东岗、奉城和辽中3个养殖场的德国镜鲤群体聚类成一个分支, 而德国镜鲤选育系与松浦群体聚类成另一分支。聚类的先后与它们在地理分布上距离远近有一定的相关性。(3)在与功能基因相关的多个微卫星基因座位上, 扩增产物呈现不同程度的缺失现象, 这些无效等位基因的产生可能与结构基因在育种中受到人工选择的影响较大有关。     

华北2蝗区东亚飞蝗种群遗传结构的比较研究

利用水平淀粉凝胶电泳对采自天津北大港和河北黄骅两个相临蝗区的东亚飞蝗（Locusta migratoria manilensis)种群进行等位酶基因频率分析,比较了这两个种群的遗传结构,等位酶酶谱分析表明,19个基因座中4个基因座（Mdh-l,Pgm,Adk,G3pd)的等位基因频率变化很小,常见等位基因的频率均高于0．95,其他基因座有2－4个等位基因,但是两个种群的等位基因频率除两个基因座（Fbp,Got-2)外都很相似,多态位点的27个χ2检验表明,由于常见等位基因纯合子的高频率的和相应杂合子的缺乏,仅有北大港种群的2个基因座（Pgi,Got-1)符合Hardy-Weinberg平衡,在每个种群内的蝗虫存在明显的遗传变异,但在种群间遗传结构极为相似,多态位点的百分数P分别为73．7％和78．9％,每个基因座的平均等位基因数A为2．9和3．1,平均每个基因座的实际杂合度几乎相等（约为0．138）,F－统计量（FST＝0．053）也表明了两个种群间的遗传 一致性,遗传相似性系数（I）高达0．938,这些结果提示,这两个种群可能属于1 个大种群,在两个种群的一定位点上的遗传多态性和分化可能都与迁飞因素有关,因为东亚飞蝗的高度扩散能力有利于遗传结构的连续分布,高度的迁飞能力也导致个体暴露于各种不同的环境,而在种群水平上的遗传为异能增强种群在各种生态条件生存和繁殖能力,因此,迁飞有利于维持东亚飞蝗种群的遗传多态性的动态平衡。     

Genetic diversity and differentiation between the two remaining populations of the critically endangered Mediterranean monk seal

The Mediterranean monk seal Monachus monachus , is a critically-endangered species of which only two populations, separated by c . 4000 km, remain: the eastern Mediterranean (150–300 individuals) and the Atlantic/western Sahara populations (100–130 individuals). We measured current levels of nuclear genetic variation at 24 microsatellite loci in 12 seals from the eastern Mediterranean and 98 seals from the western Sahara population and assessed differences between them. In both populations, genetic variation was found to be low, with mean allelic richness for the loci polymorphic in the species of 2.09 and 1.96, respectively. For most loci, the observed allele frequency distributions in both populations were discontinuous and the size ranges similar. The eastern Mediterranean population had 14 private alleles and the western Sahara had 18, but with a much larger sample size. Highly significant differences in allele frequencies between the two populations were found for 14 out of 17 loci. F _ST between the two populations was 0.578 and the estimated number of migrants per generation was 0.046, both clearly indicating substantial genetic differentiation. From a conservation perspective, these results suggest that each population may act as a source for introducing additional genetic variation into the other population.