五指山猪近交家系Ⅰ系F19～F21群体微卫星位点等位基因遗传变化

探索五指山猪近交系Ⅰ系F19～F21等位基因的遗传学规律。本研究以五指山小型猪(Susscrofa)近交系Ⅰ系53个个体为研究对象,利用14对多态性微卫星位点,通过估算其基因杂合度(H)、多态信息含量(PIC)和等位基因数(N)等参数,检测近交系Ⅰ系微卫星纯合度及等位基因遗传变化。具有多态性的14个微卫星基因座上共检测到28个等位基因,其中F19、F20和F21群体检测分别到28、26和24个等位基因,平均每个位点分别仅有2.00、1.86和1.70个等位基因;3个世代总群体平均PIC为0.24,H为0.31,均低于F18代;其中有5个基因座在第19代已经达到完全纯合。Sw1377在第21代也全部纯合。但少数基因一直表现出高度杂合状态,如Sw902、S0036和Sw874。推测这些基因可能与五指山小型猪维持其某种性状相关。本研究结果为大型动物近交系培育提供了依据。     

鳜原种群体和养殖群体遗传多样性的微卫星分析

本研究利用20对微卫星引物对鳜（Sinipelrca chuatsi）原种群体和养殖群体进行遗传多样性分析。结果表明,在鳜原种群体中检测到多态性位点14个,养殖群体11个。在两个群体中共检测到等位基因数96个,其中原种群体检测到等位基因数53个,每个位点的等位基因数在1～7之间,平均有效等位基因数为2．7390;养殖群体检测到等位基因数43个,每个位点的等位基因数在1-6之间,平均有效等位基因数为2．1284。原种群体的平均观察杂合度0．5708,Nei氏期望杂合度0．5295,平均多态信息含量PIC0．5353;养殖群体的平均观察杂合度0．3839,Nei氏期望杂合度0．4011,平均多态信息含量PIC0．5043。因此,与养殖群体相比,鳜原种群体仍有丰富的遗传多样性。本研究可为鳜种质资源的保护、监测和遗传育种提供分子水平上的数据。     

黄颡鱼微卫星标记筛选及特征分析

通过磁珠富集法首次筛选黄颡鱼微卫星标记并对其特征进行分析。用生物素标记的寡核苷酸探针(AC)8、(CT)8、(AT)7、(GATA)8、(GATT)7进行微卫星片断的富集,挑选部分阳性克隆测序获得38个含有微卫星的DNA序列,总计设计并合成了22对微卫星引物。以普通黄颡鱼基因组DNA为模板,进行PCR扩增,结果筛选出18个多态性微卫星标记,每对引物等位基因数2-8个（平均3.78）,遗传杂合度0.2225-0.8101（平均0.5755）,多态信息含量0.3425-0.7900（平均0.5336）。此外18对多态性微卫星引物中有16对在黄颡鱼属其它4种鱼类具有通用性,13对在鲇形目中的3种鱼类具有通用性。因此,可以推断筛选的多态性微卫星标记可用于黄颡鱼属和鲇形目鱼类的遗传分析。     

甘肃高山细毛羊微卫星遗传多样性及亲子鉴定研究

利用多重PCR和DNA测序技术对116只甘肃高山细毛羊(Ovis aries)13个微卫星座位的多态性进行检测,并结合父母本记录,开展亲子鉴定研究,以期为绵羊种质资源评价和亲子鉴定研究提供合理有效的微卫星座位.结果表明,甘肃高山细毛羊13个微卫星座位共检测到111个等位基因,在DRB 1-INTRO2座位检测到的等位基因数最多(14个),SRCRCP5座位最少(5个).ILSTS011座位观察杂合度最高(0.888),MAF214座位最低(0.500),除MAF214座位为中度多态外,其余12个座位均为高度多态,13个微卫星座位平均观察杂合度(Ho)为0.743,期望杂合度(He)为0.727,平均多态信息含量(PIC)为0.689.表明甘肃高山细毛羊遗传多样性较丰富.通过Cervus 2.0计算,13个微卫星座位累计父权排除概率(CPE)达到了0.999 94,11个具有高度多态的座位(除MAF214和OARJMP29外)CPE达到0.999 85,12个座位达到0.999 91,其中MAF214座位(中度多态)对CPE的影响较小,DRB1-INTRO2座位(高度多态)对CPE的影响较大,对亲子鉴定的贡献较大.本研究所分析的13个微卫星座位能够有效的提高亲子鉴定要求的精确度,可使实验操作更简便,成本降低,更适用于生产实际.     

微卫星分子标记与松浦镜鲤3个表型性状的相关分析

本文利用SPSS13.0分析软件对194尾松浦镜鲤的体重、体长和尾长值进行相关分析,并选用扩增效果好的30个微卫星标记,检测新品种松浦镜鲤繁殖群体的有效等位基因数（Ne）、观测杂合度（Ho）、期望杂合度（He）和多态信息含量（PIC）等遗传参数,以卡方检验估计群体Hardy-Weinberg平衡,最后还将30个基因座的不同基因型与个体的体重、体长和尾长进行了相关分析。结果显示,体重与体长高度线性相关,差异极显著;尾长与体重、体长分别呈显著相关,差异显著。多样性指数结果表明共检测到172个等位基因,片段长度在115～432bp之间,有效等位基因数为1.14～5.32;观测杂合度为0.07～0.92,期望杂合度为0.13～0.91;多态信息含量为0.12～0.79,其中高度多态（PIC≥0.5）21个,中度多态（0.25≤PIC≤0.5）7个,表明这个群体的多样性较高,信息含量比较丰富。我们还发现有4个遗传位点：HLJ328、HLJ693、MFW29和MFW48与体长性状显著相关（P〈0.05）,其中,位点HLJ328、HLJ693和MFW48还与体重性状显著相关（P〈0.05）,未发现与尾长性状相关的位点。本文获得的这些标记具有生长特性优势,可作为松浦镜鲤分子辅助育种的参考标记。     

白孔雀微卫星DNA遗传多样性及其分类地位

利用14对蓝孔雀(Pavo cristatus)和绿孔雀(P.muticus)的微卫星标记对白孔雀基因组DNA进行扩增,发现都能扩增出特异性条带,每对引物扩增的平均等位基因数为1.71,有7对引物具有较丰富的多态性,其中MCW0080和MCW0098标记期望杂合度分别为0.7207和0.7571,多态信息含量分别为0.658和0.695,表现出丰富的遗传多样性和较高的选择潜力。白孔雀×蓝孔雀和绿孔雀群体的遗传多样性分析结果表明,白孔雀、绿孔雀和蓝孔雀3个群体的杂合度和遗传多样性水平都很低,期望杂合度分别为0.2579、0.2482和0.2744,群体间的遗传分化系数为9.7%,群体间分化极显著(P<0.001),白孔雀与蓝孔雀的亲缘关系最近,Reynolds'遗传距离和基因流分别为0.0295和8.6112,表明白孔雀不是蓝孔雀一个亚种。     

三种笛鲷的野生群体和养殖群体遗传多样性的微卫星分析

摘要：利用微卫星标记技术,采用10对微卫星引物对南海海域红鳍笛鲷、星点笛鲷、紫红笛鲷3种笛鲷鱼的野生种群（HYE、XYE、ZYE）和养殖种群（HYA、XYA、ZYA）进行了遗传多样性分析。10对微卫星引物在6个群体中共检测到78个等位基因,每个位点的等位基因数目在1~8个之间。6个群体的观测杂合度(Ho)为0.2500~1.0000,平均观测杂合度为：ZYE(0.9550)> ZYA(0.8900),HYE(0.8950)> HYA(0.8400),XYE(0.8450)>XYA(0.8100) ;平均多态信息含量(PIC) 为0.3648~0.7964,各野生群体均大于养殖群体;三种笛鲷鱼的野生群体和养殖群体遗传距离HYE与HYA,XYE与XYA,ZYE与ZYA分别为0.1029,0.0371,0.0135,基因分化系数分别为0.0371,0.0211,0.0352。群体遗传结构分析结果表明：红鳍笛鲷、星点笛鲷和紫红笛鲷的遗传多样性较丰富,养殖群体的遗传多样性较野生群体均有一定程度的降低,野生群体和养殖群体分化较弱。     

利用微卫星标记分析边鸡遗传多样性及保种效果

利用21个微卫星标记对2个世代(0世代和1世代)的边鸡(Gallus gallus)群体的遗传结构进行检测,同时通过分析世代间遗传差异检测保种效果.结果表明,21个微卫星标记在2个世代的边鸡群体中共检测到122个等位基因,其中102个为两个世代所共有,19个为0世代所特有.0世代和1世代的边鸡群体的平均多态信息含量(PIC)分别为0.5473和0.5437;平均期望杂合度分别(He)为0.5967和0.6009;近交系数(Fis)分别为0.233和0.134.反映边鸡群体的遗传多样性较丰富,经过一个世代的选育,PIC和He两个指标维持在原有水平,而群体的近交系数有所下降,说明采用的保种方法很好地保存了边鸡的遗传多样性,同时避免了近交程度过高而导致群体出现近交衰退.     

用鲤鱼EST-SSRs分子标记分析长江黑龙江鲤种群结构

从鲤鱼ESTs的数据库（10691个）中进行微卫星序列筛选，共发现微卫星序列127个，占整个ESTs数据库的1.19%，另外从本研究室构建的鲤鱼脑组织cDNA文库中筛选到微卫星序列20个。根据筛选得到的微卫星序列设计引物68对，合成引物40对，其中27对引物能够获得预期的目的片段，20对引物在所检测的群体内及群体间表现出多态性。用此20个多态性标记分析了长江鲤和黑龙江鲤的群体遗传结构，结果表明：长江鲤和黑龙江鲤在多数位点表现出较高的遗传多样性，20个位点的平均有效等位基因数分别为3.9135、3.4820；平均观察杂合度为0.6067、0.5667；平均期望杂合度为0.6737、0.6194；平均多态信息含量为0.6308、0.5714。长江鲤的遗传结构好于黑龙江鲤，两个群体的Hardy-Weinberg偏离指数较小，群体基本保持平衡，群体结构较合理；其中HLJE1、HLJE10位点在长江鲤和黑龙江鲤的扩增结果相差较大，长江鲤的多态性明显高于黑龙江鲤，位点多态信息含量相差一倍以上，可用于群体的鉴别。     

致病疫霉基因组微卫星标记开发

致病疫霉(Phytophthora infestans)引起的马铃薯晚疫病是影响马铃薯生产的第一大真菌病害.通过致病疫霉基因组开发微卫星标记,旨为致病疫霉群体遗传结构研究提供更系统、有效的标记.利用软件SciRoKo和ClustalX对致病疫霉基因组中微卫星序列长度≥25 bp的适合标记开发的位点及两端序列进行筛选.然后在这些位点的两端序列上设计专一性PCR扩增引物,选择9株致病疫霉菌对引物的专一性和微卫星的多态性进行检测.最后利用40株不同年份和地理来源的致病疫霉菌株对可行性微卫星标记进行等位基因数的确定及其多态性评估,然后选取部分标记对40株致病疫霉群体进行遗传多样性分析.共获得了33个具多态性的微卫星标记,占开发总数的28.7％,其多态信息含量(PIC)值都在0.164～0.614之间,其中PIC≥0.5的标记有7个,0.25 ＜PIC＜0.5的标记有25个,PIC≤0.25的有1个.发现致病疫霉微卫星基因型与其交配型及地理来源有一定相关性,与甲霜灵敏感性不相关.本研究开发出一批具有多态性微卫星标记,为致病疫霉群体遗传学研究机理提供更多多态性高,稳定性强的分子标记.     

绵羊皮肤源EST-SSR标记与羊毛性状的关联性分析

为了筛选与绵羊(Ovis aries)毛性状基因座连锁更强的分子标记,本研究选择9个多态性丰富的绵羊皮肤源EST-SSR位点,检测了德国美利奴羊与中国美利奴羊级进杂交F2代群体的遗传多态性,并分析了标记与毛性状间的关联性.分析结果显示,所选的9个EST-SSR位点在供试群体中共检测到39个等位基因,多态性丰富,其中的6个位点基因型间存在显著差异;其中,与自然长度关联的位点有5个,与伸直长度关联的位点有4个,与纤维直径关联的位点有3个.6个EST-SSR位点基因型间存在显著差异;关联性标记对羊毛性状相关效应的F检验结果表明:33176918位点和33176988位点分别对羊毛自然长度、纤维平均直径具有显著的遗传效应(P＜0.05).研究结果提示,与羊毛性状相关的6个位点中,33176918号位点对羊毛的自然长度有较大的标记效应,该位点的CD基因型是羊毛自然长度的有利基因型;33176988号位点对羊毛的纤维直径有显著地标记效应,该位点的BC基因型是纤维直径的有利基因型.这两个位点可能与控制毛形状的基因座紧密连锁.     

鸭微卫星标记的筛选及其遗传多样性分析

采用磁珠富集法从AFLP片段中筛选微卫星标记,并对5个福建地方鸭品种的遗传多样性进行检测。基因组DNA用EcoRⅠ/MseⅠ内切酶酶切的同时与接头连接,酶切连接产物与用生物素标记的探针杂交,应用磁珠捕获100~2000bp含有微卫星序列的DNA片段并通过pGEM-T载体转化到大肠杆菌DH5a感受态细胞中,构建富集微卫星序列的基因组文库。随机挑选46个阳性克隆,测序获得14条含有微卫星的DNA序列并递交到GenBank（登录号：FJ599499~FJ599512）。设计合成14对微卫星引物,通过PCR优化从中选择5对引物用于5个福建地方鸭品种的遗传多样性分析。结果显示,5对微卫星引物共检测到31个等位基因,各微卫星基因座的有效等位基因数为1.969 7~2.834 4,多态信息含量和杂合度的平均值分别为0.5133和0.7480,遗传多样性丰富,说明磁珠富集法适合用于鸭微卫星标记的分离与筛选,筛选得到的5个微卫星位点可作为有效的遗传标记用于福建省地方鸭品种遗传多样性的研究。     

Variation and population structure at microsatellite and isozyme loci in wild cabbage ( Brassica oleracea L.) in Dorset (UK)

We examined variation at four isozyme and seven microsatellite loci in natural populations of Brassica oleracea L. on the coast of Dorset. All loci were polymorphic, and the diversity index of isozyme loci was similar to that of the microsatellites. Both microsatellites and isozymes showed significant spatial differentiation of genetic variation, but the scale differed between the marker types. Significant non-random mating at isozyme loci was detected within small groups of about five plants, in which adjacent individuals were separated by less than 5 m. On the other hand, non-random mating at microsatellite loci was only detected within the Dorset region as a whole. F_ST was significant for all loci and there was evidence of isolation by distance at both microsatellite and isozyme loci. From these data we infer that there is low but significant amounts of gene flow among B. oleracea populations in Dorset. The differences in the genetic structure between the two types of marker may be due to higher mutation rates at the microsatellite loci; however, the data suggest that stepwise gain or loss of single repeat units is not the principal mechanism.     

Isolation of novel microsatellite loci in Orchesella villosa (Arthropoda, Collembola)

An experimental procedure using biotin-labelled probes and streptavidin-bound magnetic beads (FIASCO) was used to produce a microsatellite-enriched library for the collembolan Orchesella villosa. PCR primers were successfully constructed for seven loci containing, respectively, five pure, one interrupted, and one compound dinucleotide microsatellite repeats. As a preliminary test of their variability, we investigated 15 individuals from 5 locations inside a dismissed mining area in southern Tuscany. All microsatellite loci showed high levels of polymorphism. The mean number of different alleles at each locus across populations was 10.1 and observed heterozygosity per locus was 0.13–0.86. Only 2 out of the 7 loci appeared to be in Hardy–Weinberg equilibrium. The potential application of these loci to test the effects of environmental contamination on the genetic structure of exposed populations is discussed.     

Genetic diversity and intra-specific phylogeny of Triticum turgidum L. subsp. dicoccon (Schrank) Thell. revealed by RFLPs and SSRs

Today, emmer wheat, T. turgidum subsp. dicoccon, widely grown in the past is a candidate crop for sustainable agriculture in Italy. As part of a research project aimed at the enhanced use of the hulled wheat germplasm, molecular characterization was carried out to understand the genetic structure of the crop and to identify accessions of interest. A collection of 194 accessions was analyzed with 15 microsatellite loci (SSRs), while only a sample of 38 accessions was tested with 19 RFLP probes. The marker loci were selected on the basis of their independent genomic distribution. Genetic distances and allelic frequencies were calculated for each marker class. The genetic relationships were visualized with dendrograms. RFLP loci were, on average, less polymorphic than SSRs. An average Dice's genetic distance of 0.22 for RFLPs vs 0.38 for SSRs was detected, while an expected average heterozygosity per locus of 0.23 for RFLPs vs 0.26 for SSRs was also estimated. With a least number of 10 loci per marker class it was possible to identify each genotype. The most diverse accessions had different geographic origins. Germplasms from Italy and Ethiopia appear to belong to a more primitive genepool, given that a group of accessions from these countries were genetically differentiated from a Russian-Iranian group.     

Genetic diversity assessment of Bulgarian durum wheat (<Emphasis Type="Italic">Triticum durum</Emphasis> Desf.) landraces and modern cultivars using microsatellite markers

The genetic diversity in a Triticum durum Desf. collection, consisting of 102 Bulgarian landraces, nine Bulgarian and 25 introduced cultivars was studied using 14 highly polymorphic microsatellite markers. A total of 100 alleles were identified, with an average of 7.14 alleles per marker. The gene diversity values (He) of the markers for the total samples ranged from 0.23 (WMS357 and WMS631) to 0.77 (WMS46), with an average of 0.52. Within the landraces that were collected from 18 sites in Southern Bulgaria showed 2–11 alleles per locus with an average of 6.07. The microsatellite analysis suggests that the genetic diversity among landraces is lower compared to the diversity levels for durum wheat in countries close to the main centers of wheat domestication. Breeding activities have caused significant reduction of the allelic polymorphism, elimination of rare alleles, and increase in the number of common alleles and the frequency of dominating alleles.     

Development and use of microsatellite markers for genetic diversity analysis of cañahua (<Emphasis Type="Italic">Chenopodium pallidicaule</Emphasis> Aellen)

Cañahua (Chenopodium pallidicaule Aellen) is a poorly studied, annual subsistence crop of the high Andes of South America. Its nutritional value (high in protein and mineral content) and ability to thrive in harsh climates make it an important regional food crop throughout the Andean region. The objectives of this study were to develop genetic markers and to quantify genetic diversity within cañahua. A set of 43 wild and cultivated cañahua genotypes and two related species (Chenopodium quinoa Willd. and Chenopodium petiolare Kunth) were evaluated for polymorphism using 192 microsatellite markers derived from random genomic cañahua sequences produced by 454 pyrosequencing of cañahua genomic DNA. Another 424 microsatellite markers from C. quinoa were also evaluated for cross-species amplification and polymorphism in cañahua. A total of 34 polymorphic microsatellite marker loci were identified which detected a total of 154 alleles with an average of 4.5 alleles per marker locus and an average heterozygosity value of 0.49. A cluster analysis, based on Nei genetic distance, clearly separated from wild cañahua genotypes from the cultivated genotypes. Within the cultivated genotypes, subclades were partitioned by AMOVA analysis into six model-based clusters, including a subclade consisting sole of erect morphotypes. The isolation by distance test displayed no significant correlation between geographic collection origin and genotypic data, suggesting that cañahua populations have moved extensively, presumably via ancient food exchange strategies among native peoples of the Andean region. The molecular markers reported here are a significant resource for ongoing efforts to characterize the extensive Bolivian and Peruvian cañahua germplasm banks, including the development of core germplasm collections needed to support emerging breeding programs.     

Cross-amplification and characterization of microsatellite loci for three species of <Emphasis Type="Italic">Theobroma</Emphasis> (Sterculiaceae) from the Brazilian Amazon

This study reports on the cross-species amplification of 23 microsatellite markers previously developed for Theobroma cacao L. (Sterculiaceae), source of chocolate in three economically important Amazonian species of Theobroma (T. grandiflorum, T. subincanum, T. sylvestre). Thirteen of the 23 microsatellite loci tested were polymorphic across the three species at 2–13 alleles per locus. The observed heterozygosity per locus varied from 0.18 to 0.84 and expected heterozygosity ranged from 0.28 to 0.87. The high level of transferability and genetic information content of these microsatellite loci indicate their usefulness for population genetic, mating system and breeding studies of these economically important Amazonian fruit trees.     

鲤鱼三、四核苷酸重复微卫星座位的筛选及特征分析*

采用磁珠富集法结合放射性同位素杂交法分离出鲤鱼(Cyprinus carpio)三、四核苷酸重复的微卫星阳性克隆1248个。对其中的384个克隆进行测序分析,共得到微卫星序列325个,其中完美型244个（75.08%）,非完美型59个（18.15%）,混合型22个（6.77%）。依据得到的微卫星序列,用Primer 3软件在线设计并合成引物145对,检测结果显示71.43%（80对/112对）的引物在野生个体间表现出多态性,等位基因数在3~12之间,多态信息含量在0.2109~0.8607之间,80%的位点处于高度多态水平。对群体的遗传结构评估结果显示,四核苷酸重复的微卫星标记在黑龙江鲤野生群体表现出的多态性水平（PIC=0.7740）高于三核苷酸重复（PIC=0.5161）和双核苷酸重复（PIC=0.49）,适用于群体间遗传差异分析和遗传连锁图谱的构建。