人工标志放流中华鲟幼鱼的降河洄游

1998— 2 0 0 2年期间 ,向长江放流人工繁殖中华鲟 2月龄稚鲟 (全长 7 5— 17 0cm) 17 5 2万尾 ,其中 7795 7尾用CWT进行标记 ;14月龄幼鲟 (全长 5 5 0— 98 0cm) 4 0 0尾 ,全部用外挂银牌和CWT双重标记。放流后沿长江及沿海收集中华鲟稚鲟和幼鲟样本 ,4年共回收稚鲟样本 6 4 0 0尾 ,幼鲟样本 13尾 ,检测到携带标记的稚鲟和幼鲟各 13尾。人工放流的幼鲟降海洄游的速度平均达到 2 8 6km/ 2 4h(7 1— 10 0 2km/ 2 4h) ,回捕时离放流点的距离从 346—2 4 5 9km ,平均 16 0 0km ,回捕的标志幼鲟有 4 6 2 %的个体来自海区。初步估算出 1999年和 2 0 0 0年人工放流个体在长江口幼鲟种群中的贡献率分别为 2 2 81%和 0 997%。结果表明 ,人工放流中华鲟稚鲟和幼鲟的生长、洄游及分布与自然种群没有明显差异。放流较大规格的幼鲟有利于提高成活率 ,而目前长江中华鲟种群的补充仍以自然繁殖种群为主。     

铲鲟微卫星引物对中华鲟的适用性研究

将21对铲鲟（Scaphirhynchus platorynchus Rafinesque)的微卫星引物在中华鲟（Acipenser sinensis Gray)基因组DNA上进行PCR扩增,14对（约占67％）引物得到了扩增产物,其中有10对（约占48％）表现为多态性,但只有2对（9．5％）引物SPl-100和Spl－168的多态性较高,且带型清晰,可直接作为分子标记应用于相关的研究,此外,对具有特异性扩增,但有Stutter band现象的4对引物的部分可分离PCR产物进行了回收和测序,并对其中的3对引物的序列进行了重新设计,最后得到两对可应用于中华鲟的引物As-100和Spl0-170b,研究结果表明,对相近种的微卫星引物进行优化设计来获得一个物种的微卫星引物是一条简捷有效的途径。     

鲸类微卫星引物对长江江豚的适用性研究

微卫星在长江江豚 (Neophocaenaphocaenoidesasiaeorientalis)中的应用研究还未见报道。本研究采用已发表的来自 6个鲸种的 2 3对微卫星引物对一个长江江豚群体DNA样本进行了微卫星扩增。结果表明其中有 7对引物在此群体中的扩增产物是稳定且多态的 ,序列分析结果表明这 7对引物的扩增产物都具有AC或GT两碱基重复单元 ,从而证明了扩增的有效性。研究结果表明用从其他鲸类分离出的微卫星引物可以快速筛选到适用于长江江豚指纹分析的引物     

湖栖鳍虾虎鱼微卫星DNA标记的 开发与群体遗传多样性分析

为研究雷州半岛湖栖鳍虾虎鱼(Gobiopterus lacustris)遗传多样性,对湖栖鳍虾虎鱼雌雄性腺进行转录组测序,采用MISA软件进行微卫星分析,共获得25 452个微卫星标记,其中包含14 708个单碱基重复类型,6 175个2碱基重复类型,3、4、5和6碱基重复类型数目分别为4 223、327、15和4。随机选取50个微卫星位点设计引物,能够扩增出清晰稳定条带的有39对,其中,11个位点表现出不同程度的多态,28个位点呈现单态。利用11个具有多态性的微卫星位点分析湖栖鳍虾虎鱼在廉江市高桥镇红树林保护区、湛江东海岛、雷州市附城镇和雷州市九龙山红树林国家湿地公园4个野生群体中的遗传多样性及遗传结构,11个微卫星位点呈现出不同程度的多态性,等位基因数(Na)2～15,平均等位基因为(6?3.9)个,有效等位基因数(Ne)、平均观测杂合度(Ho)和期望杂合度(He)分别在1.919～2.485、0.343～0.465和0.381～0.483之间,平均多态信息含量(PIC)为0.348～0.465。Hardy-Weinber平衡分析显示,4个群体的大部分位点未偏离平衡。在每个群体中进行连锁不平衡分析,有18对位点间显著(P 0.05)或者极显著(P 0.01)偏离连锁平衡。遗传分化系数在0.107～0.216之间,表明4个群体的遗传分化达到了中等水平以上。分子方差分析(AMOVA)显示,湖栖鳍虾虎鱼大部分的差异来自于群体内而非群体间。     

用微卫星标记评估红尾鲃的种群结构

本文检测了三种鲤科鱼的 1 6对微卫星引物在红尾中的适用性 ,其中 6对引物可以成功扩增 ,且 5个位点具有多态性。对采自两条不同河流的标本 ,通过检测这些多态微卫星位点的遗传变异情况 ,评估了它们在红尾种群结构分析的适合性。结果显示这 5个多态位点在上述两个样本中的平均表观杂合度分别是 0 2 93和0 4 71。这两个样本显著的基因异质性表明我们所确定的微卫星标记可用于红尾的种内遗传分化研究     

一个人工朱曦种群的遗传多态性

利用蛋白质电泳、RAPD和微卫星DNA技术,对我国濒危保护动物———人工饲养朱进行遗传多态性研究。结果表明,40只朱在前白蛋白(Pre)、白蛋白(Alb)、后白蛋白(Pa)、运铁蛋白(Tf)共4个蛋白质位点上没有发现多态性。用40个随机引物对40只朱进行RAPD分析,其中35个引物扩增出完全相同的RAPD带纹,仅筛选到5个多态性引物,朱群体的平均带纹相似率为0.86。利用其它物种的22对微卫星引物对30只朱的DNA进行检测,共筛选出4对多态性微卫星引物,扩增出13个等位基因。朱在这4对微卫星位点的平均杂合度为0.3760,平均多态信息含量为0.3382,平均等位基因数为2.075。表明朱人工饲养群体的遗传多态性比发现之初得到了一定程度的恢复,但仍然比较贫乏     

12种鲟形目鱼类mtDNA ND4L-ND4基因的序列变异及其分子系统学

鲟形目鱼类是一类在鱼类乃至脊椎动物进化史上占有很重要地位的古老濒危鱼类 ,长期悬而未决的系统演化关系为世人瞩目 .采用DNA测序技术首次测定了包括中国特有鲟形目鱼类在内的 1 2种鲟形目鱼类的mtDNA ND4L和ND4基因 ( 70 3bp)的序列 ,并进行了分子系统学分析 .从分子系统学的角度得出了如下结论 :( 1 )鳇属可以归并到鲟属 ;( 2 )达氏鲟与中华鲟的亲缘关系最近 ,很有可能为中华鲟的一陆封类型 ;( 3)环太平洋地区的鲟科鱼类可能有共同的起源 ;( 4)ND4L和ND4基因是进行鲟形目鱼类系统演化研究很好的遗传标记 .     

长江水系草鱼遗传多样性的微卫星DNA分析

利用已发表的鲤微卫星引物在草鱼中进行PCR扩增 ,结果有 5对引物 (6个座位 )能成功扩增并且有较高多态性 ,等位基因数在 3— 7个之间。这些异种扩增的草鱼微卫星符合孟德尔遗传规律。测序证明草鱼中的微卫星核心重复序列部分与鲤中的原始核心序列相似 ,也有一些变化。随后用这 6个多态微卫星座位研究了来自长江水系的四个草鱼群体的遗传结构 ,结果显示每个群体的平均等位基因数在 3 8与 4 8之间 ,平均观测杂合度 (Ho)在0 4 0 0 0与 0 5 74 1之间 ,平均期望杂合度 (HE)在 0 4 773与 0 6 4 89之间 ,有多个座位在不同的群体中偏离哈代 -温伯格平衡。遗传距离分析表明四川群体与洞庭湖群体遗传距离最远 ,而嘉鱼群体与鄱阳湖群体遗传距离较近。分子变异分析 (AMOVA)表明 ,群体内遗传变异与群体间遗传变异分别占总遗传变异的 95 6 0 %与 4 4 0 % ,固定系数 (FST)为 0 0 4 4 ,这表明长江水系草鱼目前的群体分化很微弱。     

鲤的微卫星引物对草鱼基因组分析适用性的初步研究

运用微卫星DNA-聚合酶链反应(STR—PCR)基因分型技术,选取已发表的28对鲤的微卫星引物,探讨鲤的引物用于草鱼基因组微卫星分析的可能性。通过优化PCR反应条件,消除了影子带和异源核酸双链分子两类微卫星相关假阳性带对STR—PCR分析的干扰。在此基础上,筛选出7对引物可在湘江野生草鱼基因组中扩增出特异性条带,占总数的25％;其中的4对引物(约占总数的14．3％)在8尾湘江野生草鱼小群体中即检测到了个体间等位基因的多态性。这些初步的结果表明鲤的微卫星引物可以用于草鱼基因组的分析。     

基于微卫星多重PCR技术的黄喉拟水龟亲子鉴定

利用黄喉拟水龟(Mauremys mutica)微卫星标记,筛选出16对微卫星引物,通过优化各引物比例、荧光接头浓度、退火温度和循环次数,建立了基于2组各含8个微卫星位点多重PCR体系的黄喉拟水龟亲子鉴定技术。应用2组微卫星多重PCR体系,通过ABI3130遗传分析仪以及cervus3.0软件对428只黄喉拟水龟进行了个体基因型检测和遗传多样性分析,结果显示,群体的平均等位基因数为14.190,平均多态信息含量为0.748,平均观测杂合度和期望杂合度分别为0.687、0.771。对89只候选母本及296只子代进行亲子鉴定分析,结果显示:在父本信息未知时,母本鉴定率为87%;母本获得的子代个数范围为1—12,个体间表现出巨大的差异,这为选择育种提供了物质基础。黄喉拟水龟多重PCR亲子鉴定技术的建立为群体遗传多样性分析、家系鉴定管理和选择育种提供了有效的技术手段。     

梨小食心虫微卫星标记的扩增稳定性及多态性分析

【目的】筛选适合我国梨小食心虫Grapholita molesta种群遗传学研究的微卫星位点,并依据所筛选的微卫星位点进行梨小食心虫地理种群的遗传多样性分析。【方法】利用欧洲梨小食心虫和苹果蠹蛾Cydia pomonella种群中已报道的11个微卫星位点, 分析各位点在我国12个种群257头梨小食心虫样本中的扩增稳定性,再进行其多态性分析,筛选适合的位点,然后进行种群遗传多态性分析。【结果】在分析的11个微卫星位点中, 位点Gm01, Gm03, Gm04和Cyd15无法稳定扩增; 位点Gm05扩增成功率较低, 位点Gm07遗传多态性较低; 而位点Gm02, Gm06, Gm08, Gm09和Gm10等扩增效果稳定且遗传多态性丰富。这5个稳定扩增的微卫星位点平均等位基因数量（N_A）为7.417~12.500, 平均观察杂合度（H_o）为0.366~0.655, 平均期望杂合度（H_e）为0.642~0.846, 多态信息含量（PIC）为0.800~0.935。【结论】本研究成功筛选出位点Gm02, Gm06, Gm08, Gm09和Gm10等5个微卫星位点。基于这5个微卫星位点标记的结果显示, 山东和陕西不同梨小食心虫地理种群均具有丰富的遗传多样性。 这5个位点可以适用于我国梨小食心虫种群的进一步遗传分析研究。     

Development of new microsatellite primers for green and white sturgeon

Sixty-eight primer sets for microsatellite loci were developed from microsatellite motif enriched genomic libraries of pooled DNA from the polyploid green and white sturgeon (Acipenser medirostris and A. transmontanus). Four individuals from each species were screened for polymorphism at these loci. Forty-eight loci amplified in both species, and some exhibited species-specific amplification for white or green sturgeon (8 and 12 loci, respectively). The number of alleles per locus ranged from one to 12. At least 68% of the green and 65% of the white sturgeon loci we developed are polysomic.     

从线粒体DNA控制区核苷酸序列论达式鲟、亚洲和北美洲中吻鲟的分类地位

尽管古老的鲟形目鱼类的分类及系统演化一直是中外学者感兴趣的研究课题,但迄今仍有诸多谜团未解。其中,关于亚洲远东地区和北美地区的中吻鲟(Acipenser medirostris)的分类地位争论已久。长江水系的达式鲟(A.dabryanus)和中华鲟(A.sinensis)及其它鲟属(Acipenser)鱼类之间的亲缘关系近年来也有异议。为了给上述争议提供更多的科学依据,作者测定了线粒体DNA(mtDNA)控制区 (D-loop) 的核苷酸序列,并进行了分子系统学和遗传差异分析。研究结果表明：⑴UPGMA,NJ和MP分子系统学分析方法以及遗传差异分析都支持了亚洲远东地区中吻鲟和北美中吻鲟为一个有效种的观点;⑵同样研究方法并结合相关群体遗传资料表明,长江达式鲟与中华鲟关系最近,提出了达式鲟为中华鲟的一陆封类群的假说。最后,作者认为本文提出的观点和假说还需要更多的研究来证实。     

从线粒体DNA控制区核苷酸序列论达式鲟、亚洲和北美洲中吻鲟的分类地位

尽管古老的鲟形目鱼类的分类及系统演化一直是中外学者感兴趣的研究课题 ,但迄今仍有诸多谜团未解。其中 ,关于亚洲远东地区和北美地区的中吻鲟 (Acipensermedirostris)的分类地位争论已久。长江水系的达式鲟 (A .dabryanus)和中华鲟 (A .sinensis)及其它鲟属 (Acipenser)鱼类之间的亲缘关系近年来也有异议。为了给上述争议提供更多的科学依据 ,作者测定了线粒体DNA (mtDNA)控制区(D loop) 的核苷酸序列 ,并进行了分子系统学和遗传差异分析。研究结果表明 :⑴UPGMA ,NJ和MP分子系统学分析方法以及遗传差异分析都支持了亚洲远东地区中吻鲟和北美中吻鲟为一个有效种的观点 ;⑵同样研究方法并结合相关群体遗传资料表明 ,长江达式鲟与中华鲟关系最近 ,提出了达式鲟为中华鲟的一陆封类群的假说。最后 ,作者认为本文提出的观点和假说还需要更多的研究来证实。     

Microsatellite multiplex assay for the analysis of Atlantic sturgeon populations

We have developed a multiplex assay covering 16 microsatellite loci, amplified in four polymerase chain reaction (PCR) assays, and loaded on the ABI DNA Analyzer in two separate panels. The assay was tested on 603 individuals originating from wild populations and hatchery stocks of Atlantic sturgeon. The assay was also tested on 12 individuals of European sturgeon and appeared to be almost equally useful. The multiplex assay designed in this study can be successfully applied in studies requiring high genetic resolution, such as relatedness analysis, selective breeding programs, and stock identification of Atlantic sturgeon.     

Isolation and characterization of microsatellites in Chinese soft‐shelled turtle,Pelodiscus sinensis

Fifteen polymorphic microsatellite loci were developed for the Chinese soft‐shelled turtle (Pelodiscus sinensis) from the (GT)_n microsatellite‐enriched genomic library, using the fast isolation by amplified fragment length polymorphism of sequences containing repeats protocol. The polymorphism of all 15 loci ranged from two to seven alleles with observed heterozygosities ranging from 0.03 to 0.98 (mean 0.43) in one population of 40 individuals. These novel loci will be helpful for understanding the population structure at genetic level and marker‐assisted breeding of this vulnerable species.     

Microsatellite DNA markers in Populus tremuloides.

Markers for eight new microsatellite DNA or simple sequence repeat (SSR) loci were developed and characterized in trembling aspen (Populus tremuloides) from a partial genomic library. Informativeness of these microsatellite DNA markers was examined by determining polymorphisms in 38 P. tremuloides individuals. Inheritance of selected markers was tested in progenies of controlled crosses. Six characterized SSR loci were of dinucleotide repeats (two perfect and four imperfect), and one each of trinucleotide and tetranucleotide repeats. The monomorphic SSR locus (PTR15) was of a compound imperfect dinucleotide repeat. The primers of one highly polymorphic SSR locus (PTR7) amplified two loci, and alleles could not be assigned to a specific locus. At the other six polymorphic loci, 25 alleles were detected in 38 P. tremuloides individuals; the number of alleles ranged from 2 to 7, with an average of 4.2 alleles per locus, and the observed heterozygosity ranged from 0.05 to 0.61, with an average of 0.36 per locus. The two perfect dinucleotide and one trinucleotide microsatellite DNA loci were the most informative. Microsatellite DNA variants of four SSR loci characterized previously followed a single-locus Mendelian inheritance pattern, whereas those of PTR7 from the present study showed a two-locus Mendelian inheritance pattern in controlled crosses. The microsatellite DNA markers developed and reported here could be used for assisting various genetic, breeding, biotechnology, genome mapping, conservation, and sustainable forest management programs in poplars.     

中华鲟天然群体蛋白质水平遗传多样性贫乏的初步证据

为阐明我国Ⅰ级珍稀水生保护动物中华鲟天然群体的遗传结构和遗传多样性特征,为其资源的监测量民保护提供科学依据,采用聚丙烯酰胺梯度凝胶电泳技术对中华鲟天然群体进行了蛋白质遗传多态性研究。共研究了１５种蛋白质,有４种蛋白无活呈活性很低,在有活性的１１种蛋白中人测得２６个座位;在２６个座位中,只有１个座位（ＭＤＨ－１）为多态座位。中华鲟多态座位比例（Ｐ）为３．９０％,遗传杂合度（Ｈ）为０．０４,均远远低于     

Microsatellites assessment of Chinese sturgeon (Acipenser sinensis Gray) genetic variability

Four microsatellites were used to examine the genetic variability of the spawning stocks of Chinese sturgeon, Acipenser sinensis, from the Yangtze River sampled over a 3‐year period (1999–2001). Within 60 individuals, a total of 28 alleles were detected over four polymorphic microsatellite loci. The number of alleles per locus ranged from 4 to 15, with an average allele number of 7. The number of genotypes per locus ranged from 6 to 41. The genetic diversity of four microsatellite loci varied from 0.34 to 0.67, with an average value of 0.54. For the four microsatellite loci, the deviation from the Hardy–Weinberg equilibrium was mainly due to null alleles. The mean number of alleles per locus and the mean heterozygosity were lower than the average values known for anadromous fishes. Fish were clustered according to their microsatellite characteristics using an unsupervised ‘Artificial Neural Networks’ method entitled ‘Self‐organizing Map’. The results revealed no significant genetic differentiation considering genetic distance among samples collected during different years. Lack of heterogeneity among different annual groups of spawning stocks was explained by the complex age structure (from 8 to 27 years for males and 12 to 35 years for females) of Chinese sturgeon, leading to formulate an hypothesis about the maintenance of genetic diversity and stability in long‐lived animals.     

Isolation and characterization of 16 novel microsatellite loci in two inbred strains of the Chinese hamster (Cricetulus griseus)

The Shanyi inbred A and E strains of the Chinese hamster are widely used in biomedical research, but detailed genetic characterization has been lacking. We developed microsatellite markers that could be used for genetic diversity analysis and linkage map construction. We isolated and characterized 16 novel microsatellite loci from a microsatellite-enriched genomic DNA library. These loci were genotyped in 48 animals from the two strains, and the polymorphic information content was determined. In the Shanyi A and E populations, 14 and 15 loci were found to be polymorphic, respectively, with polymorphic information content ranging from 0.1393 to 0.8082 and from 0.1109 to 0.7397, respectively. A total of 115 alleles were found for the 16 microsatellite loci in the two populations; the mean observed heterozygosity (H(O)) was 0.5191 and 0.4333 for the A and E populations, respectively, indicating marked genetic variation within the two populations. Correspondingly, the F(ST) values ranged from 0.002 to 0.9253, with an overall mean of 0.1935, indicating significant genetic difference between the two strains. The population differentiation levels were substantiated by Nei's genetic distance and full Bayesian analyses computed with STRUCTURE. Despite the genetic diversity and differentiation within and between the two inbred populations, the 48 individuals were correctly allocated into their original populations with high statistical confidence based on these 16 microsatellite loci. These novel microsatellite loci should be useful genetic markers for these two Chinese hamster inbred strains.