基于叶绿体单倍型的山东丹参居群遗传结构分析

依据叶绿体psbA-trnH基因间隔区的序列多态对来自山东地区丹参的3个自然居群和1个栽培居群进行了遗传结构研究.结果显示,80个供试材料中共有10种单倍型,核苷酸多样性为0.024,叶绿体DNA单倍型进化分支分析把10种单倍型分为2大支,但每支都由4个居群的单倍型共同构成;Nei多样性指数为0.284,Shannon信息指数为0.406,多态位点百分比率为70%;4个居群总基因多样性为0.905,居群内平均基因多样性指数为0.747,遗传分化系数为0.175;不同居群间的遗传分化系数为0.017～0.199,基因流为2.013～28.912;AMOVA分析揭示居群间只有5%的遗传差异.研究结果表明,山东丹参遗传多样性水平较高,但由于彼此较高的基因交流,居群间遗传分化水平低,仅有的遗传差异主要存在于居群内;另外,序列谱系关系分析表明山东栽培丹参来源复杂且具有丰富的遗传多样性.     

伏牛山区连翘遗传多样性研究

利用2个叶绿体及1个核基因片段,对伏牛山区3个连翘野生种群30个个体进行了遗传多样性分析.结果表明,叶绿体基因数据得到4个单倍型(H1～H4),单倍型多样性指数(h)为0.545±0.066,核酸多样性指数π为0.001 72±0.000 21.核基因数据得到10个单倍型(A～J),单倍型多样性指数为0.501±0.076,核酸多样性指数π为0.002 53±0.000 49.核基因种群间遗传分化(ФST)为0.167,小于叶绿体基因ФST(0.886),种群间基因流(Nm)为1.250,显著大于叶绿体基因Nm(0.030).研究发现,伏牛山区野生连翘种群目前仍具有较高的遗传多样性,但部分人类活动较多的地区已经造成了连翘遗传多样性的降低.连翘种群间地理距离会限制种子介导的基因流,但在小区域范围内对花粉所介导的基因流影响不明显.     

基于核基因LEAFY的中国珍稀濒危植物中华水韭的遗传多样性分析

利用核基因LEAFY第二个内含子片段对中国现存中华水韭（Isoetes sinensis Palmer）的遗传多样性进行分析,探讨了中华水韭自然居群的遗传多样性结构及其形成机制。结果显示：现存中华水韭7个自然居群共105个样本中存在78个单倍型,单倍型多样性（H_d）为0.989,核苷酸多样性（π）为0.021,遗传差异主要存在于居群内（72%）,且存在较高的基因流（N_m=0.59）。同时,居群遗传学分析结果发现中华水韭居群不存在明显的谱系地理格局（G_ST>N_ST）;Mantel检验中Rxy值为-0.286,P（rxy-rand≥rxy-data）值为0.370,表明居群遗传距离和地理距离之间没有明显相关性;UPGMA聚类分析显示处于海拔较高位置的2个居群与其它5个居群遗传关系较远;中性检验（Taijima's D、Fu & Li's D^*和F^*）检测结果均为负值,基于稳定模型的失配分布检测显示为多峰。根据中华水韭居群的地理位置,推测中华水韭的遗传结构可能与水系、海拔分布及其杂交后代多倍化的物种形成过程相关。     

异叶泽兰的遗传多样性和居群历史动态研究

异叶泽兰属于菊科泽兰属,是该属分布海拔相对较高的植物,分布在青藏高原东部和横断山海拔1700~3000 m的地区。该文利用ycf6-psbM和rpl32-trnL两个叶绿体DNA(cpDNA)片段以及核DNA片段ITS(nITS)作为分子标记,研究了异叶泽兰的遗传多样性及其分布特征,探讨了其居群历史动态。结果表明:(1)叶绿体片段联合分析结果显示,异叶泽兰在物种水平遗传多样性水平不高,单倍型多态性指数(H_d)为0.656,核苷酸多态性(π)为0.00161;而ITS的基因型多态性指数(H d)为0.687,核苷酸多态性(π)为0.00235。(2)cpDNA和nITS分析结果均显示,异叶泽兰居群水平总的遗传多样性大于居群内平均遗传多样性,遗传变异主要发生在居群间,居群间存在明显的遗传分化(cpDNA:G_st=0.679,N_st=0.655,F st=0.655;nITS:G_st=0.543,N_st=0.370,F_st=0.584),但是由于N_st值小于G_st值,因此异叶泽兰的分布不具有明显的谱系地理结构。(3)基于单倍型地理分布以及Network分析推测,横断山区南部(川西南-滇西北)和云南中部可能是异叶泽兰在第四纪冰期时的两个避难所,中性检验和失配分析的结果支持异叶泽兰在冰期后未发生过居群扩张。     

黑藻cpDNA分子标记的筛选及其遗传多样性的初步研究

本研究从18个叶绿体DNA（cpDNA）分子标记中筛选出3个有效分子标记trnL-trnF、trnH-psbA和RPL16,利用它们对中国黑藻（Hydrilla verticillata（L.f.）Royle）8个居群进行了遗传多样性研究。结果显示,黑藻8个居群40个样本共有12个单倍型,总变异位点数为135个;单倍型多样性指数为0.9064,遗传多样性主要存在于居群间（89.6%）。其中单倍型Hap4同时存在于地理距离相隔较远的陕西汉中（SXHZ）和浙江余姚（YY）居群中。研究结果表明黑藻可以通过无性繁殖的方式快速完成居群扩张,同时,地理隔离和生境差异可能导致居群间基因流较低,奠基者效应和遗传漂变可能对黑藻的遗传结构产生重要影响。     

利用rp132-trnL分析阿尔金山自然保护区真藓居群遗传多样性

本研究对分布在中国新疆维吾尔自治区阿尔金山自然保护区的6个真藓(Bryum argenteum)居群的遗传结构及遗传多样性进行比较。通过对32条叶绿体DNA的rpl32-trnL序列的核苷酸序列变异的分析,发现了14种单倍型,存在201个可变位点;分子变异分析显示有51.02%的遗传变异发生在居群间水平,居群内部的遗传变异为48.98%,真藓居群间的遗传分化程度较高略大于居群内的遗传分化。居群遗传变异的分化系数为0.510 2,基因流值为0.48,显示各居群间的基因流低。单倍型多态性水平为(0.780 2±0.076 0),核苷酸多态性水平为(0.058 59±0.020 09),表明真藓居群遗传多样性丰富。对所有变异位点进行的Taijma's检验的结果是Taijma's D值为-1.567 05 (p0.10),显示所有变异符合中性进化假说。     

两种濒危水韭植物迁地保护居群的基因流动态及回归重建保育遗传管理策略

采用微卫星分子标记对中华水韭（Isoetessinensis）安徽休宁、浙江建德和东方水韭（I.orientalis）浙江松阳三个孑遗居群的迁地保护居群开展了遗传多样性检测与遗传结构分析。7对多态性微卫星引物在36个迁地保护亚居群的720个样本中共检测到59个等位基因,每位点平均等位基因数（A）为8·43。迁地保护亚居群均维持很高的遗传多样性,多态信息含量（PIC）平均为0·707。迁地保护亚居群间遗传分化较低,遗传分化系数GST仅为0·070,居群间具有较大基因流（Nm=3·59）。单因素方差分析发现水韭孢子或孢子体在沿主要水流方向上的长距离传播能力要强于弱水流方向上的短距离传播能力,水流动态对水韭植物的基因流有重要影响。这与UPGMA聚类分析中迁地保护亚居群按邻近位置或水流相通程度优先聚类的结果相一致,水流所带动的强大基因流导致了不同孑遗居群来源的迁地保护亚居群间的遗传混杂。建议在开展水韭植物的迁地保护或回归自然重建时,对具有地方适应分化或者显著性进化的水韭植物居群应相互隔离而不宜配置在一起,以避免远交衰退的遗传风险。     

濒危植物宽叶羌活天然居群cpDNA非编码区多态性分析

采用叶绿体DNA非编码区直接测序的方法,对青海、甘肃、四川3个省区内濒危植物宽叶羌活14个天然居群的遗传多样性和遗传结构进行了研究,以明确其遗传背景,为宽叶羌活的保护提供理论依据。结果表明:(1)14个居群138个个体的序列长度介于509~515bp;碱基组成A+T含量较高(平均67.6%)。(2)根据序列的核苷酸变异共鉴定出31个单倍型,其中9种单倍型(H5、H1、H9、H10、H16、H17、H19、H20、H22)为居群所共享,而且单倍型H5分布最广、在10个群体的67个个体中检测到,占总样品数的48.56%。(3)14个居群宽叶羌活具有高水平单倍型多样性(Hd=0.750 3)和较高水平核苷酸多样性(Pi=0.007 11),但31个单倍型没有按地理分布形成明显的族群,各地理单元中的单倍型相互混杂,没有明显的地理分化模式。(4)AMOVA分析、居群间分化度(FST=0.602 4)分析和基因流(Nm=0.330)分析的结果一致,表明宽叶羌活大部分遗传变异(60.24%)发生在居群间,居群间的基因流较低,群体间变异是宽叶羌活的主要变异来源。(5)14个居群的遗传距离在0.000~0.007,平均为0.003,表明居群间的亲缘关系较远,且居群间的遗传距离与地理距离之间无显著相关关系(P=0.143),说明宽叶羌活居群遗传变异的分布没有明显的地理趋势。研究认为,宽叶羌活居群间高的遗传变异可能是由基因流受阻、遗传漂变、地理隔离造成的,并提出了对该物种遗传多样性的保护策略。     

若尔盖湿地青海沙蜥红原亚种线粒体ND4-tRNAleu基因的序列变异和遗传多样性

研究了青海沙蜥红原亚种(Phrynocephalus vlangalii hongyuanensis)分布于四川若尔盖辖曼(XM)、红原(HY)和甘肃玛曲(MQ)这3个地理单元7个居群的ND4-tRNAleu基因的序列变异和遗传多样性。72个序列比对得到785bp的片断,含变异位点7个,定义了9种单倍型。结果显示7个居群总的核苷酸多样性较低(0.00231±0.00016),单倍型多样性较高(0.806±0.024);就单个居群而言,MQa、MQb、XMb的遗传多样性低,XMc相对较高。各居群间的Kimura双参数遗传距离小(0.001—0.005),最大遗传距离出现在MQa和XMa之间,XM单元各居群之间遗传距离最小。但分子变异分析(AMOVA)显示3个单元间出现了显著差异(P<0.01),遗传变异主要存在于地理单元间,占62.61%。除MQ单元形成一支,单倍型网络图没有显示出单倍型和地理位置的对应关系,XM各居群以及HY混杂在一起。XM单元单倍型的不配对分布(Mismatch distribution)为明显左移的单峰,同时Fu’sFstest得到负值(Fs=-2.21937),这都暗示XM单元可能经历了近期种群扩张,这是其单倍型多样性较高和核苷酸多样性较低的原因。MQ单元遗传多样性低而与其他单元显著分化,推测与黄河在若尔盖玛曲之间贯通有关。近期沼泽的形成对XMb的隔离时间短,使得其遗传多样性低但还不足以形成大的遗传差异。     

青藏高原地区山生柳遗传多样性研究

利用叶绿体非编码区片段研究分布于青藏高原地区的山生柳居群遗传多样性,对未来山生柳生态环境和青藏高原地区物种丰富度的保护具有指导意义。该研究设计并筛选出cpDNA引物5'trnG2G-3'trnG(UUC)和5'rpS12-rpL20,用扩增出的片段和对应的联合片段进行后续的遗传多样性分析。结果表明:通过山生柳的联合片段检测到35种单倍型,单倍型多态性0.626,核苷酸多态性0.000 85。中性检验Tajima’s D(-2.286 70,P0.01)和Fu’s Fs(-5.298 05,P0.02)都是显著负值,推测山生柳个体数近期经历过扩张。AMOVA分析显示,居群内和居群间遗传变异分别为93.70%和6.30%,表明居群内的变异是山生柳遗传变异的主要来源。居群间遗传分化程度中等偏低(FST=0.063),基因流(N_m)为7.439,说明山生柳各居群的基因交流非常频繁,不同地理居群间存在一定的基因流动。遗传分化系数N_(ST)(0.075)大于GST(0.068)和基于遗传距离和单倍型的UPGMA聚类分析,表明山生柳12个居群分为4组且与居群的地理分布没有明显相关性。山生柳是进行有性繁殖还是无性繁殖主要受环境因素的影响,居群内变异是山生柳遗传变异性的主要来源,居群间基因交流频繁。     

Phylogeography of Camellia taliensis (Theaceae) inferred from chloroplast and nuclear DNA: insights into evolutionary history and conservation

ABSTRACT: BACKGROUND: As one of the most important but seriously endangered wild relatives of the cultivated tea, Camellia taliensis harbors valuable gene resources for tea tree improvement in the future. The knowledge of genetic variation and population structure may provide insights into evolutionary history and germplasm conservation of the species. RESULTS: Here, we sampled 21 natural populations from the species' range in China and performed the phylogeography of C. taliensis by using the nuclear PAL gene fragment and chloroplast rpl32-trnL intergenic spacer. Levels of haplotype diversity and nucleotide diversity detected at rpl32-trnL (h = 0.841; pi = 0.00314) were almost as high as at PAL (h = 0.836; pi = 0.00417). Significant chloroplast DNA population subdivision was detected (GST = 0.988; NST = 0.989), suggesting fairly high genetic differentiation and low levels of recurrent gene flow through seeds among populations. Nested clade phylogeographic analysis of chlorotypes suggests that population genetic structure in C. taliensis has been affected by habitat fragmentation in the past. However, the detection of a moderate nrDNA population subdivision (GST = 0.222; NST = 0.301) provided the evidence of efficient pollen-mediated gene flow among populations and significant phylogeographical structure (NST > GST; P < 0.01). The analysis of PAL haplotypes indicates that phylogeographical pattern of nrDNA haplotypes might be caused by restricted gene flow with isolation by distance, which was also supported by Mantel's test of nrDNA haplotypes (r = 0.234, P < 0.001). We found that chlorotype C1 was fixed in seven populations of Lancang River Region, implying that the Lancang River might have provided a corridor for the long-distance dispersal of the species. CONCLUSIONS: We found that C. taliensis showed fairly high genetic differentiation resulting from restricted gene flow and habitat fragmentation. This phylogeographical study gives us deep insights into population structure of the species and conservation strategies for germplasm sampling and developing in situ conservation of natural populations.     

Molecular phylogeography and conservation genetics of Sladenia celastrifolia inferred from chloroplast DNA sequence variation

The genetic variation and structure of Sladenia celastrifolia Kurz, a species of conservation concern, were investigated. Analyses of two chloroplast DNA loci (trnS-trnGand atpB-rbcL intergenic regions) were carried out for 24 populations of S. celastrifolia and five haplotypes were identified. High levels of genetic differentiation (G_ST = 1, F_ST = 1) were detected, which may be a result of limited gene flow caused by geographic isolation. Analysis of molecular variance suggests that the existence of marked phylogeographical structure within the haplotype distribution is probably due to geographic barriers among populations. The haplotype network and mismatch distribution analyses did not detect any signals for recent population expansions in S. celastrifolia. Thus, it can be inferred that the species likely persistedin situ during climatic oscillations. Considering its genetic diversity and uniqueness, conservation strategies are further discussed for this species.     

Phylogeography and genetic structure of disjunct Salix arbutifolia populations in Japan

Disjunct geographic distributions of boreal plant species at the southern edges of their ranges are expected to result in low genetic diversity and high genetic differentiation in the disjunct populations. This prediction was tested in a riparian willow, Salix arbutifolia, distributed in the northeastern Eurasian continent and the Sakhalin, Hokkaido, and Honshu Islands, using chloroplast DNA haplotypes and nuclear microsatellite genotypes. Hokkaido and Honshu populations shared a chloroplast haplotype identical to a closely related species, S. cardiophylla. This haplotype was divergent from haplotypes in the Eurasian continent (Primorsky) and the Sakhalin Island. In the nuclear microsatellites, most Hokkaido populations were genetically closer to Primorsky populations than to Sakhalin populations in spite of the geographical vicinity between Sakhalin and Hokkaido. The unexpected genetic divergence between Sakhalin and Hokkaido implies a complicated history of migration and colonization. The most peripheral populations in Honshu had the lowest genetic diversity and were most differentiated from the others. Thus, low genetic diversity and high genetic differentiation at the range periphery suggest substantial effects of genetic drift on genetic structure in the disjunct populations of Salix arbutifolia at the southern edge of its range.     

Chloroplast DNA variation and genetic structure of Miscanthus sinensis in southwest China

Miscanthus sinensis is a dominant perennial C4 grass with the potential to being a feedstock crop in North America, Europe, and China. Variation in chloroplast DNA sequence was used to obtain information regarding the genetic diversity and structure of populations of M. sinensis in southwest China. Chloroplast DNA, trnL-F and rpl20-rps12 sequences from seventy-five individuals representing 14 populations of M. sinensis were used to study the sequence variation. Seven haplotypes and 16 polymorphic sites (2.7%) were identified. Five substitutions, 6 indels, and 5 existing substitutions and indels sites, were detected through splicing these two gene segments. The genetic diversity within the studied populations (diversity of haploids, h = 0.561, nucleotide diversity, π = 0.00504) was low, this may be affected by the relatively larger effect of genetic drift on the chloroplast DNA, reflecting smaller effective populations than nuclear DNA. Genetic variance within the populations was higher than that between the populations, suggesting that higher gene flow may exist within these populations. The results of parsimony network in seven haplotypes indicated that H1 and H2 may be ancient haplotypes, which may help guide future research on the origin of M. sinensis. Our results provide information on the genetic diversity and structure of M. sinensis and may assist future studies on the phylogeography of M. sinensis.     

Genetic diversity in endangered Notopterygium forbesii Boissieu based on intraspecies sequence variation of chloroplast DNA and implications for conservation

A thorough understanding of the levels and partitioning of genetic variation across populations and geographical regions of endangered species is a prerequisite to ensure effective conservation and/or restoration activities. Here, we examined chloroplast DNA (cpDNA) trnH-psbA intergenic spacer sequences variation within Notopterygium forbesii, an endangered and endemic perennial herb in China. Sequence data obtained from 141 individuals in 14 populations revealed twenty-two haplotypes. A high level of haplotype diversity (Hd = 0.81) and low level of nucleotide diversity (Pi = 0.0047) were detected. Low genetic differentiation among populations and also among regions was consistently indicated by both hierarchical analyses of molecular variance (AMOVA) and the structure of a neighbor-joining tree. Low level of population differentiation between populations or between regions in cpDNA sequences may be due to effects of the abundance of ancestral haplotype sharing and the high number of private haplotypes fixed for each population. Based on our results, we proposed some conservation strategies.     

青藏高原东北部及其邻近地区无苞香蒲的谱系地理学

第四纪冰期气候的反复变化对青藏高原及邻近地区植物的种群地理分布及种群遗传结构产生了巨大的影响。本研究对青藏高原东北部及其邻近地区无苞香蒲(Typha laxmannii)的15个种群148个个体的叶绿体rpl32-trnL间隔区和核基因(植物螯合肽合成酶, PS)进行测序, 共发现2个叶绿体单倍型和8个核基因单倍型。所有的单倍型被共享, 高原种群没有特有的单倍型。邻近地区种群的叶绿体遗传多样性和核基因遗传多样性分别是高原种群的4倍和2倍。高原种群的遗传分化水平明显高于邻近地区种群, 其中高原种群的遗传分化主要存在于东部种群与西部种群之间。研究结果表明, 冰期后从多个避难所回迁至高原台面和由此产生的奠基者效应造成了无苞香蒲在青藏高原东北及邻近地区目前的遗传多样性和基因谱系地理分布格局。     

Genetic structure of the endangered Leucomeris decora (Asteraceae) in China inferred from chloroplast and nuclear DNA markers

The genetic variation and structure of Leucomeris decora, an endangered species in China were investigated. Analyses of three chloroplast DNA (cpDNA) regions (the rpl16 intron, trnQ-5′rps16 intergenic spacer and rpl32-trnL intergenic spacer) and one nuclear gene (GAPDH: encoding glyceraldehyde 3-phosphate dehydrogenase) were conducted on 11 L. decora populations. Low levels of cpDNA genetic diversity were found in this species and within populations, with the identification of 2 haplotypes in a total of 2,745 bp, while the level of genetic diversity revealed by the nuclear gene GAPDH was relatively high, indicating that random losses of genetic polymorphisms from populations may have occurred recently. High levels of genetic differentiation among populations for both markers were detected in L. decora, which could be a consequence of the limited gene flow caused by geographic isolation among populations. An analysis of molecular variance revealed at the nuclear locus suggested the presence of geographic structure within the haplotype distribution possibly due to geographical barriers among populations. The haplotype network and mismatch distribution analyses did not detect the signal for a recent population expansion in L. decora. L. decora may persist in situ during climatic oscillations. Based on the genetic diversity and uniqueness of the populations, conservation strategies are discussed for this endangered species.     

Chloroplast DNA phylogeography of the endangered Japanese red maple (Acer pycnanthum): the spatial configuration of wetlands shapes genetic diversity

Aim Japanese red maple (Acer pycnanthum K. Koch) is an endangered species which grows in discrete wetland ecosystems within a limited geographical range. It is an important relic of geologic time, an endemic of unique wetland ecosystems and an indicator of hotspots of plant species diversity. However, information on its genetic characteristics across its range is lacking. Our aim was to determine the genetic structure and diversity of the species and make recommendations for conservation. Location Wetlands in central Honshu Island, Japan. Methods We collected leaf samples from 400 individuals of A. pycnanthum in 30 populations, extracted total DNA from each and sequenced three non‐coding regions of chloroplast DNA. Results We identified nine haplotypes. High haplotype diversity (0.81) and the occurrence of rare haplotypes in eight distant populations suggest that wetlands provided multiple, adequate‐size refuges during the Last Glacial Maximum. We found only one to four haplotypes within populations. The high degree of differentiation (G_ST = 0.83) implies that gene flow by seeds among populations is restricted. Eight populations demonstrated a positive contribution to the total genetic diversity owing to occurrence of rare and private haplotypes. Such populations are concentrated in the south‐western part of the species distribution. According to the spatial autocorrelation analysis, there were significant spatial clusters of populations, which were characterized by similar haplotype composition. Using the haplotype distribution, samova and barrier detected nearly identical genetic boundaries. Main conclusion In spite of the species’ limited geographical range, we identified a relatively high number of haplotypes and a clear geographical structure. We propose six management units, which can be used for future conservation activities, such as introduction of new individuals for on‐site conservation projects and seed collection for ex situ conservation.     

Restricted gene flow in fragmented populations of a wind-pollinated tree

Fragmentation of natural populations can have negative effects at the genetic level, thus threatening their evolutionary potential. Many of the negative genetic impacts of population fragmentation can be ameliorated by gene flow and it has been suggested that in wind-pollinated tree species, high or even increased levels of gene flow are a feature of fragmented populations, although several studies have disputed this. We have used a combination of nuclear microsatellites and allele-specific PCR (AS-PCR) analysis of chloroplast single nucleotide polymorphisms (SNPs) to examine the levels and patterns of genetic diversity and population differentiation in fragmented populations of juniper (Juniperus communis) in Ireland and inform conservation programs for the species. Significant population differentiation was found for both chloroplast and nuclear markers, indicating restricted gene flow, particularly over larger geographic scales. For conservation purposes, the existence of genetically distinct clusters and geographically localised chloroplast haplotypes suggests that the concept of provenance should be taken into account when formulating augmentation or reintroduction strategies. Furthermore, the potential lack of seed dispersal and seedling establishment means that ex-situ approaches to seed and seedling management may have to be considered.