Development and characterization of eleven polymorphic microsatellite loci from South China field mouse (Apodemus draco)

One population distributed in Yunnan of China was regarded as a new species based on mitochondrial cytochrome b sequences. However, the usefulness of mitochondrial sequence data in determining species boundaries is not universally agreed upon, the frequency data from multiple nuclear gene loci is necessary in determining species boundaries. So, we describe in this paper the isolation and characterization of eleven microsatellite loci in the South China field mouse from genomic DNA-enriched libraries. The eleven loci were tested in 24 individuals from two populations in Southwest China. These loci were highly polymorphic with numbers of alleles per locus ranging from 9 to 24 and expected heterozygosities from 0.898 to 0.967. Eight loci followed Hardy–Weinberg expectations after Bonferroni correction for multiple comparisons. No significant linkage association was found among all these loci. The eleven polymorphic microsatellite loci will be useful in determining species boundaries of the South China field mouse.     

华东地区青冈种群的等位酶变异

华东地区６个青岗种群等位酶变异的检测结果表明,青冈种群的遗传变异较大,每位点含有的等位基因较多,平均为２．３个,种水平的平均每位点等位基因数目为２．４。种群水平有效等位基因数目为１．４４６,种水平为１．４６７。Ｈａｒｄｙ－Ｗｅｉｎｂｅｒｇ平衡检验表明,青冈种群中不少位点偏离该平衡,其原因是种群内含有较多的纯合子,表现为多数位点的固定指数大于０,其中有９个位点的固定指数与０的偏差达到显著水平。     

Isolation and characterization of microsatellite loci in the western long-fingered bat, Miniopterus magnater

We isolated and characterized 10 microsatellite loci in the western long-fingered bat, Miniopterus magnater. These loci were tested on 48 individuals from Anhui Province of China, and all loci were highly polymorphic. The mean number of observed alleles per locus was 13.6 (range from six to 27). Observed and expected heterozygosity values ranged from 0.364 to 0.957, and from 0.676 to 0.951, respectively. After Bonferroni correction, four loci deviated significantly from Hardy-Weinberg equilibrium. No pairs of loci were in linkage disequilibrium. These polymorphic markers will be used to examine population structure and genetic diversity in this species.     

Isolation and characterization of microsatellite loci in the long-fingered bat Miniopterus fuliginosus

We isolated and characterized 10 microsatellite loci in the long-fingered bat Miniopterus fuliginosus. These loci were tested on 48 individuals from Anhui Province of China, and all loci were highly polymorphic. The mean number of observed alleles per locus was 13.6 (range from six to 27). Observed and expected heterozygosity values ranged from 0.364 to 0.957, and from 0.676 to 0.951, respectively. After Bonferroni correction, four loci deviated significantly from Hardy-Weinberg equilibrium. No pairs of loci were in linkage disequilibrium. These polymorphic markers will be used to examine population structure and genetic diversity in this species.     

Development and characterization of microsatellites in Torreya jackii (Taxaceae), an endangered species in China

? Premise of the study: Polymorphic microsatellite loci were developed in Torreya jackii, an endangered species in China, to provide markers for further studies on the genetic diversity of this species. ? Methods and Results: Eight polymorphic loci and one monomorphic locus were developed and characterized in four T. jackii populations (Xianju, Songyang, Pujiang, and Tonglu) from Zhejiang Province, China. The number of alleles per locus ranged from one to eight across 80 T. jackii individuals. At the eight polymorphic loci, the observed heterozygosity ranged from 0.150 to 1.000 and the expected heterozygosity ranged from 0.185 to 0.796. ? Conclusions: The microsatellite loci developed and characterized in this study will facilitate future analyses of the genetic diversity of T. jackii. Such information will aid in designing strategies to conserve this currently endangered species.     

Isolation and characterization of polymorphic microsatellite loci in Wuchang bream (Megalobrama amblycephala)

Wuchang bream (Megalobrama amblycephala) is an economically important fish in China. From a (GT)₁₃‐enriched genomic library, 20 microsatellites were developed. Nine of these 20 loci were polymorphic in a test population with allele numbers ranging from two to four, and the observed and expected heterozygosities ranging from 0.2609 to 0.7826 and from 0.3739 to 0.7546, respectively. In the cross‐species amplifications, six of these nine loci were also polymorphic in white amur bream (Parabramis pekinensis). These polymorphic microsatellite loci are potentially useful for population genetics of Wuchang bream and its closely related species.     

Genetic Diversity and Population Structure for the Conservation of Giant Spiny Frog(Quasipaa spinosa) Using Microsatellite Loci and Mitochondrial DNA

The giant spiny frog(Quasipaa spinosa) is an endangered species with a relatively small distribution limited to southern China and Northern Vietnam. This species is becoming increasingly threatened because of over-exploitation and habitat degradation. This study provides data on the genetic diversity and population genetic structure of the giant spiny frog to facilitate the further development of effective conservation recommendations for this economically important but threatened species. We examined 10 species-specific microsatellite loci and Cyt b genes(562 bp) collected from 13 wild populations across the entire range of this species. Results of 10 microsatellite loci analysis showed a generally high level of genetic diversity. Moreover, the genetic differentiation among all 12 populations was moderate to large(overall F_(ST) = 0.1057). A total of 51 haplotypes were identified for Cyt b, which suggests high haplotype nucleotide diversities. Phylogeographic and population structure analyses using both DNA markers suggested that the wild giant spiny frog can be divided into four distinct major clades, i.e., Northern Vietnam, Western China, Central China, and Eastern China. The clades with significant genetic divergence are reproductively isolated, as evidenced by a high number of private alleles and strong incidence of failed amplification in microsatellite loci. Our research, coupled with other studies, suggests that Q. spinosa might be a species complex within which no detectable morphological variation has been revealed. The four phylogenetic clades and some subclades with distinct geographical distribution should be regarded as independent management units for conservation purposes.     

Isolation and characterization of polymorphic microsatellite markers in lesser kestrel (Falco naumanni) and cross-amplification in common kestrel (Falco tinnunculus)

Lesser kestrel, Falco naumanni, is a colonial and migratory species breeding in part of the Mediterranean Basin and part of central Asia and north-east of China and Mongolia. This species is catalogued in IUCN red list category as vulnerable. Twenty microsatellite loci were selected from libraries enriched in AC or AG tandem repeats and specific PCR were devised from their flanking sequences. Most microsatellites (14) were found polymorphic among 30 individuals of F. naumanni representing 20 reproduction areas of the species in the region of Extremadura, Spain. Polymorphisms were detected by size variation of the amplified loci, which allele number and observed heterozygosity ranged from 3 to 20 and from 0.300 to 0.933, respectively. Cross-species amplification showed that 13 of selected loci were also found polymorphic in common kestrel species (Falco tinnunculus). Novel polymorphic microsatellites will serve to conservation studies in lesser kestrel.     

Isolation and characterization of microsatellite loci in the flat‐headed bat (Tylonycteris pachypus)

Microsatellite loci were developed in the flat‐headed bat (Tylonycteris pachypus) from genomic DNA using an enriched library method. Nine loci were tested on 48 individuals sampled from Guangxi Province, China. The mean number of observed alleles per locus was 6.4 (range 4–12). Observed and expected heterozygosity values ranged from 0.24 to 0.83 and from 0.30 to 0.89, respectively. One locus revealed significant departure from Hardy–Weinberg equilibrium and no significant linkage disequilibrium was detected between loci pairs. These markers will be used to examine genetic structure and parentage analysis in this species.     

Development of twelve polymorphic microsatellite loci in polyploid endangered Omphalogramma vincaeflora Franch. (Primulaceae)

A total of twelve polymorphic microsatellite loci were developed from polyploid endangered species, Omphalogramma vincaeflora (Primulaceae). These loci were screened for variability among 45 individuals from three populations in China. The primers amplified loci with allele number ranging from 3 to 9, with an average of 4.25 per locus. Polymorphism information content ranged from 0.23 to 0.86. Nei’s genetic diversity ranged from 0.34 to 0.86. These primers provide an opportunity to use polymorphic DNA markers to study the population genetic structure and its breeding system in this species.     

厚壳桂新微卫星体分子标记的开发与应用

生境破碎化是导致全球生物多样性危机的主要原因,即使常见物种也会受到很大影响.厚壳桂(Cryptocarya chinensis)是我国南亚热带季风常绿阔叶林群落演替顶极物种,由于这一森林植被的破坏而呈片断化分布.在以往的研究中,我们利用筛选到的一些微卫星体对其遗传多样性研究后发现其可能以无性生殖为主.为进一步证实这一发...     

Microsatellite development for the endangered Yangtze sturgeon (Acipenser dabryanus Duméril, 1869) using 454 sequencing

The Yangtze sturgeon (Acipenser dabryanus) is an endemic species in China. Using 454 sequencing, eight polymorphic tri‐, tetra‐, penta‐, and hexanucleotide microsatellite loci were isolated in this study. The raw sequence data from a one‐eighth run of 454 sequencings were 38.0 Mbp containing 94 222 reads/sequences. Of 80 microsatellite loci, only eight loci were polymorphic in a population of 30 individuals. The number of alleles per locus ranged from 4 to 14 (mean 7.62), and the observed heterozygosities varied between 0.46 and 0.88 (mean 0.74). Cross amplification was tested in congeneric species Acipenser sturio and Acipenser sinensis. These new microsatellite markers will be useful for further studies on genetic variation, parentage analysis, and conservation management for this critically endangered species.     

马尾松和黄山松两个核基因位点的群体遗传多样性和种间分化

利用两个核基因座位C3H和GI, 对重叠分布于中国东南部的两个松属(Pinus)物种马尾松(P. massoniana)和黄山松(P. hwangshanensis)的22个群体88个个体进行了遗传多样性和种间分化模式研究。在这两个核基因座位上, 两种植物都表现出较低的核苷酸多样性水平(马尾松π_sil = 0.001 71; 黄山松π_sil = 0.003 40), 但是马尾松要显著低于黄山松; 在种内分化水平上, 马尾松的种内遗传分化也明显低于黄山松(马尾松F_ST = 0.059; 黄山松F_ST = 0.339)。这可能是由于黄山松的海拔分布高于马尾松, 而高海拔分布使黄山松的分布区域更加片段化, 促使其形成较高的种内遗传多样性和遗传分化。分子变异分析(AMOVA)发现, 两物种基于两个核基因座位的种间差异为48.86%, 而GI基因座位上的种间差异明显高于C3H座位(GI: 77.24%, C3H: 20.48%), 同时, 基因谱系显示两物种的共享单倍型仅在C3H座位上存在。结合这两个基因的功能, 推测GI基因可能在物种形成过程中受到了一定的选择压力, 因为GI基因参与调控植物的开花时间, 而C3H与木质素表达水平的调控有关。不同的选择压力使得GI的进化速度相对较快, 从而加速了黄山松和马尾松的物种分化。     

广西区西南桦天然居群遗传多样性的研究

 以采自广西区11个西南桦(Betula alnoides)天然居群的种子培育出的幼苗为材料,取其嫩叶开展21种酶系统的水平切片淀粉凝胶电泳实验,运用作者改进的Tris-马来酸提取缓冲液 (含30%PVP 40 000和1%2-巯基乙醇),筛选出AMP(氨基肽酶Aminopeptidase)、FBA(果糖二磷酸酶Fructose-bisphosphate aldolase)、GDH(谷氨酸脱氢酶Glutamate dehydrogenase)、G6PD(6-磷酸葡萄糖脱氢酶Glucose-6-phosphate dehydrogenase)、IDH(异柠檬酸脱氢酶Isocitrate dehydrogenase)、MDH(苹果酸脱氢酶Malate dehydrogenase)、PGD(6-磷酸葡萄糖酸脱氢酶Phosphogluconate dehydrogenase)、PGI(磷酸葡萄糖异构酶Phosphoglucoisomerase)、PGM(磷酸葡萄糖变位酶Phosphoglucomutase)和SKD(莽草酸脱氢酶Shikimate dehydrogenase)等10种酶,获得了清晰且可重复的酶谱。通过谱带遗传分析确定了15个位点,其中有6个单态位点,9个多态位点 (0.95标准),具40个等位基因。在居群水平上,西南桦的多态位点百分数 (P) 为55.2%,平均每个位点的等位基因数 (A) 为2.00,平均预期杂合度 (He) 为0.204。均超过Hamrick (1992)等提出的远交风媒木本植物的平均值(53.0%,184%和0154),表明西南桦的遗传变异水平高。在11个西南桦居群内,实际杂合度 (Ho)均高于预期杂合度,出现杂合子过量,可能存在利于杂合子的自然选择。西南桦遗传多样性与地理位置相关不显著。居群2 (靖西地州)、5 (平果海城)、9 (田林者苗) 包含绝大部分的等位基因,而且具较高的遗传多样性,应加以保护和管理,作为其基因资源就地保存的基地。本研究解决了以西南桦嫩叶为材料进行等位酶分析的关键技术,为进一步开展西南桦乃至桦木属树种的遗传结构和遗传多样性等提供了技术基础;同时掌握了西南桦天然居群的遗传多样性现状,为其有效保护和合理经营以及西南桦的遗传改良提供了理论依据。     

Characterization of nine novel microsatellite loci for the Venus clam (Cyclina sinensis)

The Venus clam, Cyclina sinensis, is one of the most important bivalves in China marine aquaculture. Using (CA)(15)-enriched genomic libraries of this species, nine novel polymorphic microsatellite loci were isolated and characterized. The mean number of observed alleles per locus was 16 (range 8-24). The observed and expected heterozygosity ranged from 0.119 to 0.872 and from 0.626 to 0.931, respectively. Three loci had significant departure from Hardy-Weinberg equilibrium and non-significant linkage disequilibrium was found among all nine loci. These highly informative microsatellite markers should be useful for population genetic analyses of C. sinensis.     

南方红豆杉紫杉烷高含量植株系RAPD初步研究

南方红豆杉（Ｔａｘｕｓｃｈｉｎｅｎｓｉｓｖａｒ． ｍａｉｒｅｉ（Ｌｅｍｅｅ ｅｔＬｅｖｌ．） Ｃｈｅｎｇ ｅｔＬ．Ｋ．Ｆｕ） 是我国提取抗癌药紫杉醇的原料植物之一,但分布于不同地区的南方红豆杉其紫杉醇含量差异很大。应用ＲＡＰＤ技术,用２１ 个引物共扩增２５６ 条条带,其中有２２４ 条有多态性（８７．８ ％） 。结合紫杉烷及紫杉醇含量测定和植物形态分类,初步探讨了紫杉烷高含量植株的分子特征、形态和生境的相互关系。在供试的１２ 个地区２４ 个样品中,提出３ 个地区存在紫杉烷高含量植株系的可能性,为资源利用和保护以及优良品种的培育提供基     

Rapid isolation and characterization of polymorphic microsatellite loci in the mud crab, Scylla paramamosain (Portunidae)

Scylla paramamosain is a widespread and commercially important species of coastal marine crab. We identified 13 polymorphic microsatellite loci from a genome library constructed with 5'-anchored PCR method. Thirty-two S. paramamosain from the East China Sea were used to analyze the characteristics of these loci. The number of alleles per locus ranged from 3 to 8, with a mean of 5.923. Observed and expected heterozygosities ranged from 0.500 to 0.875 and from 0.500 to 0.859, respectively. Eleven of the 13 loci were highly polymorphic (polymorphic information content >0.5). All of the 13 novel loci were in Hardy-Weinberg equilibrium after Bonferroni's correction (P < 0.0038). There was no null allele, stuttering errors or evidence of allelic dropout in any of the loci analyzed by MICRO-CHECKER. According to pairwise tests, no significant linkage disequilibrium was found among the 13 loci (P < 0.0038, adjusted value). These novel developed microsatellites will be useful for studies of genetic variation, population structure, conservation genetics, and molecular-assisted selective breeding of S. paramamosain.     

Isolation and characterization of 13 microsatellite loci in Rhinolophus pusillus (least horseshoe bat) with cross-amplification in five related species

We used the enriched genomic library method to isolate and characterize dinucleotide microsatellite loci in the least horseshoe bat, Rhinolophus pusillus. Seventeen loci were obtained and tested on 31 individuals sampled from Guangxi Province in southern China. Thirteen of these markers were polymorphic with expected heterozygosity ranging from 0.821 to 0.909. A total of 164 alleles were detected and the number of alleles per locus ranged from 9 to 16 (mean 12.6). These polymorphic markers will be used to assess population structure in R. pusillus. In addition, successful cross-amplification in five congeneric bat species suggests most of these markers will also be useful for studying related species.     

RAPD Analysis on High-taxane-contained Accessions of Taxus chinensis var. mairei

Taxus chinensis var. mairei (kemee et Levi. ) Cheng et L.K. Fu is one of the resource plants in China. The taxot contents from different localities are variable. RAPD and HPLC analysis were carried out with 24 samples from 12_localities. A total of 256 loci were amplified among which 224 loci were polymorphic (87.8%). Based on the HPLC and morphological analysis, the relationship among RAPD markers, morphology and habitat was explored. According to the results, high-taxane-contained accessions are present in three localities. This study might be of benefit to the resource conservation and species improvement.     

Isolation and characterization of 18 polymorphic microsatellite markers for the “Female Ginseng” Angelica sinensis (Apiaceae) and cross-species amplification

Angelica sinensis (Apiaceae) is an endangered alpine herb that is widely used as a medicinal plant in traditional Chinese medicine (TCM). Wild populations of A. sinensis have become quite rare in China. Thus, population genetics studies of this species are urgently needed for its effective conservation and sustainable use. However, to date, no microsatellite loci have been isolated in A. sinensis. To address this issue, we isolated 18 polymorphic loci and genotyped 120 individuals collected from 6 populations. The number of alleles per locus ranged from 1.2 to 5.5, and the average was 2.4. The observed and expected heterozygosity per locus for a population varied, respectively, from 0.000 to 0.983 (averaged at 0.198) and from 0.066 to 0.661 (averaged at 0.333). Deviation from the Hardy–Weinberg equilibrium (p < 0.01) was observed for 4 to 14 loci in various populations. These microsatellite markers were cross-amplified in 10 species affinis, and 7 loci were successfully amplified in all species. These microsatellite markers are useful for genetic studies, the conservation management of A. sinensis, and identification of A. sinensis.