Development and diversity of microsatellite markers for endangered species Clinostigma savoryanum

A set of microsatellite markers was developed for Clinostigma savoryanum, an endemic palm species distributed in the Bonin Islands. We obtained 233 sequences that were unique, containing microsatellites from an enriched library. Twelve loci were screened for their feasibility to be used as high resolution genetic markers using each 30 individuals from two insular populations, Haha-jima and Mukou-jima. They showed polymorphisms of two to eight alleles per locus and expected heterozygosities of 0.124–0.789. There is no evidence for significant scoring error due to stuttering, large allele dropout and null alleles at 95% confidence interval except for the presence of null alleles in CLS00-77 of Mukou-jima population.     

Development of SCAR markers distinguishing pure seedlings of the endangered species Morus boninensis from M. boninensis × M. acidosa hybrids for conservation in Bonin (Ogasawara) Islands

To support the recovery of Morusboninensis Koidz. (Moraceae), a criticallyendangered species distributed in the BoninIslands of Japan, we need to distinguishbetween pure M. boninensis and hybrid(M. boninensis × M. acidosa Griff.)seedlings within populations derived from M. boninensis mother trees. Therefore, wedeveloped three co-dominant SCAR (SequenceCharacterized Amplified Region) markers todistinguish between M. boninensis, M. acidosa and their hybrids. All threemarkers could clearly identify the species andthe hybrids, although we observed one case of aputative null allele or a backcross at onelocus (Mos0251-1). Hence, we suggest that pureand hybrid seedlings could be unambiguouslyidentified by the simultaneous use of the threemarkers. When we investigated a seedlingpopulation produced by a single mother treegrowing in the Iguma-bay population ofHaha-jima Island, the proportion of embryosformed by pollination from M. acidosapollen was about 13%. All hybrid seedlingsidentified according to the molecular markersshowed incisions in their leaves, amorphological character that has never beenobserved in pure M. boninensisindividuals. Our results suggest that M.boninensis can potentially be multiply byexcluding hybrids from the seedling pools,using incisions on the leaves as morphologicalmarkers to identify them.     

Genetic diversity and conservation evaluation of a critically endangered endemic maple,Acer yangbiense,analyzed using microsatellite markers

The newly discovered endemic maple Acer yangbiense of China has only five individuals left in the wild, and thus has been classified as a plant species with extremely small populations (PSESP). PSESP species call for emergency protection procedures, such as ex-situ conservation and reintroduction. Our objectives were to examine the genetic diversity of A. yangbiense and to evaluate former conservation strategies from a genetic point of view. Our results suggested that A. yangbiense was not genetically depauperate, but its genetic loss at a species level was obvious. A parentage analysis indicated a high selfing-rate in A. yangbiense and suggested the existence of a previously unknown wild individual. Former conservation strategies did not include all genetic variations of the wild population, and gene diversity of the ex-situ conserved seedlings is lower than that of the wild population. From our findings, we make suggestions to guide the subsequent protection of this species.     

Development of microsatellite markers for the endangered Neotropical tree species Tibouchina papyrus (Melastomataceae)

We isolated and characterized 12 microsatellite loci for Tibouchina papyrus (Melastomataceae), an endangered species with narrow and disjunct range, endemics to a few localities in "cerrado rupestre" from Central Brazil. These microsatellites were obtained by sequencing of a genomic shotgun library for primer design. Leaves from 96 individuals collected in the three known local populations were genotyped using the 12 primers designed to analyze the polymorphisms at each locus. The number of alleles per locus ranged from one to six; two loci were monomorphic. Among the polymorphic loci, expected heterozygosities ranged from 0.161 to 0.714. Combined paternity exclusion probability was 0.957 and combined genetic identity (0.051) was high for studies on parentage. Tibouchina papyrus is a rare and endemic tree species of outcrop quartzite and sandstone soils, with highly isolated populations, which may have lead to the low degree of polymorphism that we detected. Also, motifs of most loci are larger than dinucleotide, which typically display lower levels of polymorphism.     

DNA分子标记技术在濒危物种保护中的应用

近20年来,随着分子生物学技术的迅猛发展,涌现出一批高效、可靠的DNA分子标记技术.本文论述了限制性片段长度多态性、微卫星DNA、随机扩增多态性DNA、扩增片段长度多态性等DNA分子标记技术的基本原理及技术特点;同时,介绍了DNA分子标记在濒危物种种群遗传学研究、致危因素分析及保护策略的制定等保护生物学方面的应用.     

微卫星分子标记在濒危动物保护遗传学研究中的应用

微卫星DNA广泛分布于真核生物基因组中,具有多态性高、共显性遗传、选择中性、易于操作等特点,是一种极具应用价值的分子遗传标记,近年来在濒危动物保护遗传学研究中得到越来越多的应用。微卫星DNA高度多态性提供的高分辨率遗传信启,使其不仅适合个体水平的亲子鉴定与交配系统研究,而且也已成为种群遗传结构与多样性分析的有效分子标记。微卫星分析所需的DNA量极少,用非损伤性方法获取的极少量样品或陈旧样品就能用于有效分析,方便了濒危动物野外调查工作的开展,并且可以利用年代久远的馆藏历史标本揭示种群的重要历史进程。另外,某些微卫星DNA大小在近缘物种间可相互区分,这使得部分物种的DNA分子鉴别将更为简便。但微卫星分子标记的座位筛选和特异引物开发耗时费力,一定程度上限制了其广泛应用。针对不同的研究目的选择合适的分子标记方法将有助于更好的揭示问题本质。     

Development of microsatellite markers for Quercus miyagii Koidz. (Fagaceae), an endemic species in the Ryukyu Islands, Japan

Quercus miyagii is an endemic tree species in the Ryukyu Islands, Japan. We isolated and characterized 15 microsatellite loci in this species. The number of alleles ranged from 2 to 16 and expected heterozygosities from 0.07 to 0.92. This set of markers is potentially useful to investigate the genetic structure, gene flow, and the biogeographic history of Q. miyagii in the Ryukyu Islands, Japan.     

Polymorphic microsatellite markers for endangered spotted suslik,Spermophilus suslicus

Spotted suslik Spermophilus suslicus is one of the most endangered mammal species in Poland. Over the last 50 years, it lost more than 90% of its populations and about 70% of its individuals. In order to establish a conservation and reintroduction program, the knowledge of population structure of the species is crucial. We have developed nine polymorphic microsatellite loci to analyse the population structure. Six of the primer sets also amplify polymorphic markers in the European suslik Spermophilus citellus.     

Genetic diversity assessment and ex situ conservation strategy of the endangered Dendrobium officinale (Orchidaceae) using new trinucleotide microsatellite markers

Dendrobium officinale (Orchidaceae) is an endangered plant species with important medicinal value. To evaluate the effectiveness of ex situ collection of D. officinale genetic diversity, we developed 15 polymorphic trinucleotide microsatellite loci of D. officinale to examine the genetic diversity and structure of three D. officinale germplasm collections comprising 120 individuals from its germplasm collection base and their respective wild populations consisting of 62 individuals from three provinces in China. The three germplasm collections showed reductions in gene diversity and average number of alleles per locus, but an increase in average number of rare alleles (frequency?≤?0.05) per locus in comparison to their wild populations. However, the differences in gene diversity between the germplasm collections and wild populations were not statistically significant. The analysis using STRUCTURE revealed evident differences in genetic composition between each germplasm collection and its wild population, probably because the D. officinale individuals with distinct genotypes in each wild population were unevenly selected for establishing its germplasm collection. For conservation management plans, we propose that D. officinale individuals with rare alleles need to be conserved with top priority, and those individuals with the most common alleles also should be concerned. The 15 new microsatellite loci may be used as a powerful tool for further evaluation and conservation of the genetic diversity of D. officinale germplasm resources.     

濒危物种巴东木莲的等位酶遗传多样性及其保护策略

为了确定中国特有濒危植物巴东木莲(Manglietia patungensis)的就地保护优先单元和制定迁地保护的取样策略,采用超薄平板聚丙烯酰胺等电聚焦电泳技术对巴东木莲的7个野生居群的等位酶遗传多样性及遗传结构进行了分析。通过对8个酶系统19个酶位点的分析结果来看,巴东木莲具有较高的遗传多样性和一定程度的遗传分化。每位点平均等位基因数（A）为1.57,多态位点百分率（P）为48.1%,平均预期杂合度（H_e）为0.192。巴东木莲居群间的遗传分化系数（G_ST）为0.165,说明其16.5％的遗传变异存在于居群间;基于遗传分化系数计算的基因流（N_m）为1.27,说明居群间存在适度的基因流;固定指数(F)为-0.191,显示巴东木莲居群杂合体轻微过量,纯合体略显不足。本研究表明,目前巴东木莲仍具有较高的遗传多样性并且在其适宜生境中可以完成自然更新,因此应采取以就地保护为主的综合保育策略。     

Development of microsatellite markers for Polemonium kiushianum (Polemoniaceae), a critically endangered grassland plant species in Japan

Eight microsatellite loci were isolated and characterized for Polemonium kiushianum (Polemoniaceae), an endangered perennial herb endemic to the Kyushu region in Japan. The number of individuals of this species is presumed to be about 400 individuals. The loci showed polymorphisms with two to ten alleles per locus. The expected and observed heterozygositis ranged from 0.039 to 0.744 and 0.039 to 0.784, respectively. Although significant deviations (P < 0.05) from Hardy–Weinberg equilibrium were detected for one locus, no significant deviations were detected for other seven loci. There was no evidence of significant linkage disequilibrium between any two of the loci. The markers described here will be useful to establish conservation programs based on genetic traits of P. kiushianum.     

Development and characterization of microsatellite markers for endangered species Stipa pennata (Poaceae) and their usefulness in intraspecific delimitation

Molecular Biology Reports - Stipa pennata (Poaceae), has become a rare and endangered species in Central Europe due habitat loss and fragmentation. This species is characterized by high...     

Development and characterization of microsatellite markers for the endangered anchialine squat lobster Munidopsis polymorpha

Species of the genus Munidopsis are typically distributed in bathyal and abyssal zones, but the anchialine species Munidopsis polymorpha is an exception. It inhabits a volcanic tube on Lanzarote Island (Canary Islands, NE Atlantic) and is currently listed as endangered due to its highly restricted distribution and degree of endemism. Microsatellite loci were isolated from partial genomic libraries that had been enriched for AC, ACAG, GATA, AAAC and AAG repeat sequences. Eight loci were polymorphic in a sample of 24 individuals. The number of alleles per locus ranged from 2 to 4 with observed and expected heterozygosities ranging from 0.083 to 0.875 and from 0.080 to 0.681, respectively. These markers will be used to evaluate levels of genetic diversity and inbreeding, providing essential information for the development of a management and conservation strategy for this species.     

浙江省珍稀濒危植物物种多样性保护的关键区域

浙江省植物资源丰富,有野生维管植物215科1196属3283种,其中包含了我国特有属49个,浙江省特有植物约200余种,许多物种十分稀少,并受到严重威胁,亟待保护。但浙江省需优先保护的植物物种的分布并不是均匀的,有些地区的重要保护对象比较集中,对这些地区我们应该给予更多的保护。本文根据维管植物物种多样性、珍稀濒危植物的物种数量及个体数量特征、受威胁状况以及浙江省特有植物的分布情况,提出了浙江省珍稀濒危植物保护的7个关键区域：以西天目山为中心的浙西北山区,以古田山为中心的浙西山区,以九龙山为中心的浙西南山区,以凤阳山－百山祖为中心的浙南山区,以括苍山为中心的浙东山区,以天台山为中心的浙东丘陵,以普陀山为中心的舟山群岛。最后提出了浙江省植物物种多样性保护的几点建议。     

Development and characterization of microsatellite markers for two dioecious Ficus species

Microsatellite markers for Ficus montana and Ficus septica were developed using genomic libraries enriched for di‐, tri‐ and tetranucleotide repeats. The subsets of five and three best scorable primer pairs were characterized on 24 F. montana and 36 F. septica individuals, respectively. For F. montana, loci showed five to 14 alleles per locus and expected heterozygosities ranged between 0.23 and 0.87. For F. septica, loci showed three to five alleles per locus and expected heterozygosities ranged between 0.36 and 0.49. Four primer pairs (two from each subset) cross‐amplified in the other species, indicating transportability of the markers within the genus Ficus.     

遗传多样性与濒危植物保护生物学研究进展

尽管对于濒危物种的遗传学人们已经进行了大量研究,但是种群遗传学在植物保护中的实际地位尚存在很大争议。濒危物种的遗传多样性可能会由于遗传漂变、近交的作用而丧失;但这种丧失更可能是濒危的结果而不是濒危的起因。遗传多样性水平与物种生存力之间没有任何必然的联系。但植物种群遗传结构如果由于自交不亲和等位基因的丧失和与亲缘种杂交造成的遗传同化而发生改变,那么它对物种生存力会产生明显负作用。     

Development of microsatellite markers in Dendrobium fimbriatum Hook, an endangered Chinese endemic herb

As an endangered endemic herb, Dendrobium fimbriatum, is under threat from numerous impacts. In order to analyse the genetic diversity and structure of this endangered species, we provide details of 10 microsatellite loci (out of 15 primer pairs designed) which showed polymorphic for D. fimbriatum. These loci were used to screen 25 individuals from across the species' geographical range. Ten loci were polymorphic with 2 to 19 alleles; three loci were monomorphic, while the rest produced no amplification fragments. These loci will be used to investigate population genetic structure, genetic diversity, conservation, and individual authentication in the endangered D. fimbriatum.