云南古茶园栽培大叶茶和大理茶群体的叶绿体RPL32-TRNL核苷酸变异和遗传分化

对云南地区栽培的大叶茶（Camellia sinensis var. assamica）9个居群和大理茶（C.taliensis）3个居群的遗传多样性和居群遗传结构进行了比较研究。通过对94条叶绿体DNA RPL32-TRNL序列的核苷酸序列变异进行分析,共发现了7个单倍型。研究结果表明,大叶茶（h=0.728,π=0.00469）比大理茶（h=0.610,π=0.00225）拥有更为丰富的遗传变异,但其居群遗传分化水平（Gst=0.580,FST=0.612）却低于大理茶（Gst=0.696,FST=0.773）。AMOVA分析进一步证实了它们的遗传变异主要存在于居群间,且大叶茶（61.21%）低于大理茶（77.34%）。相对于大理茶,遗传多样性水平在本研究所取的大叶茶居群间存在着较大的差异,单倍型多态性水平变化范围为0~0.809,核苷酸多态性水平变化范围为0~0.00487。最后,讨论并提出了科学有效地保护我国古茶园茶种资源的建议和对策。     

核基因组微卫星标记揭示大理茶参与了普洱茶的驯化过程

利用11个核基因组微卫星标记对普洱茶3个居群、大理茶3个居群及过渡型大理茶2个居群共104株古茶树进行了遗传学分析。研究表明,普洱茶、大理茶和过渡型大理茶居群的遗传多样性相对较低,平均等位基因(Na)为4.852,平均香农多样性指数(I)为1.17,平均期望杂合度(He)和观测杂合度(Ho)分别为0.59和0.52,其中大理茶的遗传多样性水平低于普洱茶和过渡型大理茶。AMOVA分析表明,普洱茶和大理茶之间遗传分化显著(FST=0.305),遗传变异主要在居群内(分别为93.51%和89.41%),而居群间的遗传变异较低(分别为6.49%和10.59%)。主成分分析和STRUCTURE聚类分析均支持大理茶和普洱茶为不同的组,过渡型大理茶主要由大理茶驯化而来,并在栽培过程中与大理茶产生了遗传分化。在混栽的大理茶和普洱茶居群间存在由大理茶向普洱茶的明显基因渐渗,证实了大理茶参与了普洱茶的驯化过程。最后,讨论并提出了对大理茶和普洱茶古茶树资源保护的相关建议。     

中国山茶属4种2变种核型研究

本文采用去壁低渗法研究了我国山茶属植物4种2变种的核型。根据Levan等的命名系统,各个种的核型可简式为大理茶2n=20m+2m(SAT)+8sm;勐腊茶2n=20m+2m(SAT)+6sm+2sm(SAT);德宏茶2n=20m+8sm+2sm(SAT);苦茶2n=20m+9sm+1sm(SAT);茶2n=18m+2m(SAT)+12sm;白毛茶2n=18m十2m(SAT)+9sm+1sm(SAT)。这些种都是二倍体种(2n=2x=30),未发现多倍体。在勐腊茶核型中发现2个超数染色体(B-chromosome)。核型的不对称性表明,这些种均属于Stebbins核型分类的2A型核型。这些种在“随体数目和位置,最长染色体与最短染色体之比,臂比大于2:1的染色体比例,着丝点端化值,染色体绝对长度”方面的变异是清楚易见的。核型的变异表明,这些种的染色体进化顺序为大理茶—→勐腊茶—→德宏茶—→普洱茶—→白毛茶—→苦茶—→茶。这一结果与张宏达提出的山茶属植物的分类系统基木吻合。本文还讨论了山茶属植物核型的杂合性和多态性。本文中勐腊茶、德宏茶、苦茶的染色体数目和核型及大理茶的核型为首次报道。     

Chemical Analysis of Old Tea Trees in Bai Ying Shan Mountain and the Origin of Cultivated Tea

The contents of dalichasu (1,2-di-O-galloyl-4,6-O-(S) hexahydroxydiphenoyl-β-D-glucopyranose), catechins, theogallin, gallic acid, caffeine and theanine in old tea trees from Bai Ying Shan mountain were quantitatively analyzed by HPLC method, while their total polysaccharides and polyphenols were measured by an UV Vis spectrophotometer. Based on the chemical composition and morphological variation, the germplasm resource diversity of tea trees in Bai Ying Shan Mountain and their relationship with wild Camellia taliensis and cultivated tea (Csinensis var assamica) were discussed. The results provided not only the scientific data of quality evaluation of the old tea trees in Bai Ying Shan Mountain, but also the chemical evidence for the origin of cultivated tea and the hypothesis that Ctaliensis might be one of the origins of cultivated tea during the long natural variation and artificial breeding.     

HPLC Analysis of Theanine and Gallic Acid inthe Species of Camellia sect. Thea

Bymeans of HPLC technique, the theanine and gallic acid in the leaves of six species and three varieties of Camellia sect. Thea were analyzed. The results showed that both of them were generally occured in these plants and the contents in C. taliensis and C. kwangsiensis are mostly close to those of C. sinensis var. assamica. Since the leaves of C. taliensis have also been historically used for preparing tea by folk peoples of Yunnan province, it could be suggest that C. taliensis might one of the original plants of tea     

ISSR and SRAP Markers Reveal Genetic Diversity and Population Structure of an Endangered Slipper Orchid,Paphiopedilum micranthum (Orchidaceae)

Paphiopedilum micranthum is an endangered pink slipper orchid mainly distributed in the limestone areas of southwestern China. Wild populations of this species have been seriously threatened by excessive collections, rampant smuggling for export, and habitat destruction. We used 15 ISSR markers and 11 SRAP markers to investigate the genetic diversity and structure of 15 natural populations. A high degree of diversity was observed at the species level (ISSR: PPB=9166%, He=03839; SRAP: PPB=9929%, He=02806). Certain degree of genetic differentiation among populations (ISSR: Gst=02577; SRAP: Gst=02383) was detected maybe caused by low gene flow (ISSR: Nm=07201; SRAP: Nm=07991). Consistent with the results of Principal Coordinate Analysis, the UPGMA dendrogram analysis divided the 15 populations into two main clades. In addition to geographic distance, the difference in elevation was another natural factor contributing to this differentiation. Knowledge about genetic diversity and structure gained from our study will be beneficial for the development of reasonable and efficient strategies to conserve this endangered species.     

中日5个岛屿山茶种群遗传多样性研究

运用ISSR分子标记法对中、日两国5个岛屿天然山茶种群共150个个体进行遗传多样性分析。结果表明:筛选出的20条引物扩增得到205条清晰条带,其中183条为多态性条带,多态位点百分比(PPB)为89.27%。经POPGENE软件分析,山茶种群平均多态位点百分比(PPB)为72.00%,Nei’s基因多样性指数(HE)为0.2743,Shannon信息多态性指数(H)为0.4023,种群水平遗传多样性较高。基因分化系数Gst=0.2033,表明遗传变异主要存在于种群内个体间。Mantel检验(r=0.7989,P<0.05)和UPGMA聚类表明岛屿地理隔离对山茶种群遗传分化具有重要影响。基于岛屿山茶种群遗传结构的分析,建议加强我国岛屿自然种群的就地保护力度。     

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.     

Population genetics and phylogeography of endangered Oxytropis campestris var. chartacea and relatives: arctic-alpine disjuncts in eastern North America

Fassett's locoweed (Oxytropis campestris var. chartacea, Fabaceae) is an endangered perennial endemic to Wisconsin. Patterns of genetic variation within and among six remaining populations and their relationship to other members of the O. campestris complex were analysed using AFLPs from 140 accessions across northern North America. Within-population measures of genetic diversity were high (mean expected heterozygosity HE = 0.16; mean nucleotide diversity pi = 0.015) compared with other herbaceous plants. Estimates of among-population differentiation were low (FST = 0.12; PhiST = 0.29), consistent with outcrossing. Genetic and geographical distances between populations were significantly correlated within Fassett's locoweed (r2 = 0.73, P < 0.002 for Mantel test) and O. campestris as a whole (r2 = 0.63, P < 0.0001). Individual and population-based phylogenetic analyses showed that Fassett's locoweed is monophyletic and sister to O. campestris var. johannensis. Morphometric analyses revealed significant differences between Fassett's locoweed and populations of var. johannensis. The first chromosome count for Fassett's locoweed indicates that it is tetraploid (2n = 32), unlike hexaploid var. johannensis. High within-population diversity and relatively low among-population differentiation are consistent with populations of Fassett's locoweed being relicts of a more continuous Pleistocene distribution. Our data support the continued recognition of Fassett's locoweed and protection under federal and state regulations. High levels of genetic diversity within populations suggest that maintain-ing the ecological conditions that favour the life cycle of this plant may be a more pressing concern than the erosion of genetic variation.     

白莺山古茶的化学成分分析与栽培茶树的起源

应用高压液相色谱(HPLC)技术,对白莺山古茶的大理茶素、儿茶素类、茶掊素、没食子酸、咖啡因和茶氨酸的含量进行分析.同时,应用分光光度法测定茶多糖和茶多酚含量.在与野生大理茶及栽培大叶茶(普洱生茶)进行多成分比较的基础上,结合形态性状的变异与进化,讨论白莺山古茶种质资源的多样性与野生大理茶和栽培大叶茶的相互关系.分析研究结果不仅为白莺山古茶的品质评价提供可靠的科学数据,为白莺山丰富的古茶种质资源的深人系统研究和合理开发利用提供基础,同时,通过多种过渡类型的化学成分分析与比较,为栽培大叶茶的起源和大理茶作为大叶茶的野生基源之一的假说提供了植物化学方面的证据.     

大别山山核桃天然群体遗传结构的初步分析

利用RAPD分子标记技术检测了大别山山核桃3个天然居群的遗传多样性和遗传结构。20条10 bp随机引物共检测到238个扩增位点,其中多态性位点162个,多态位点百分率(PPB)为68.1%。居群水平Shannon’s多态性信息指数(I)介于0.2651~0.2801之间;居群水平Ne i’s基因多样性指数(H)介于0.1789~0.1890之间。遗传变异计算显示大别山山核桃居群间基因分化系数(Gst)为0.4063,分子方差分析(AMOVA)表明居群间基因分化水平为0.4177,居群间基因流(Nm)为0.7306,说明大别山山核桃大部分变异存在于居群内,居群间基因交流相对较少。这一结果符合大别山山核桃风媒、异交的繁育系统特点,但其居群间基因分化程度明显高于异交植物的平均水平(Gst=0.1930)。地理隔离、居群内近交及居群间基因流受阻可能是形成目前大别山山核桃天然群体遗传结构的主要因素。     

Low genetic diversity and high genetic differentiation in the critically endangered Omphalogramma souliei (Primulaceae):implications for its conservation

Omphalogramma souliei Franch. Is an endangered perennial herb only distributed in alpine areas of SW China. ISSR markers were applied to determine the genetic variation and genetic structure of 60 individuals of three populations of O. Souliei in NW Yunnan, China. The genetic diversity at the species level is low with P= 42.5% (percentage of polymorphic bands) and Hsp=0.1762 (total genetic diversity). However, a high level of genetic differentiation among populations was detected based on different measures (Nei's genetic diversity analysis: Gst=0.6038; AMOVA analysis: Fst=0.6797). Low level of genetic diversity within populations and significant genetic differentiation among populations might be due to the mixed mating system in which xenog-amy predominated and autogamy played an assistant role in O. Souliei. The genetic drift due to small population size and limited current gene flow also resulted in significant genetic differentiation. The assessment of genetic variation and differentiation of the endangered species provides important information for conservation on a genetic basis. Conservation strategies for this rare endemic species are proposed.     

海南粗榧(Cephalotaxus mannii Hk.f.) 5个种群遗传多样性及分化研究

采用垂直板型聚丙烯酰胺凝胶电泳测定了海南粗榧5个种群遗传多样性和遗分化程度。结果表明：海南粗榧种群遗传多样性水平较低,多态位点比率P＝0．33,等位基因平均数A＝1．33,平均期望杂合度为He=0．135,观察杂合度为Ho＝0139。种群间遗传分化程度较低,基因分化系数Gst=0.123,种群间遗传一致度和遗传距离的均值分别为0．9719、9．0288。黎母山种群与其它种群分化最大,其原因可能与传粉时盛行的风向有关。     

中国特有植物卧龙沙棘自然群体的RAPD分析

应用12个随机引物对卧龙沙棘全部2居群共28个个体进行了RAPD分析。结果表明,卧龙沙棘具有较丰富的遗传多样性,多态位点百分率为78.05%,Nei’s基因多样性h=0.2553,Shannon’s多态性信息指数Ι=0.3841。分布范围狭窄的卧龙沙棘在亚种水平的遗传多样性明显高于分布较广的中国沙棘。在居群水平上,卧龙沙棘同样具有很高的遗传多样性,平均多态位点百分率为63.42%,Nei’s基因多样性h=0.2193,Shannon’s多态性信息指数I=0.3287。卧龙沙棘的基因分化系数Gst=0.1425,表明遗传变异主要存在于居群内。AMOVA的结果也表明,在全部的遗传变异中,19.18%的遗传变异存在于居群之间,与Gst值基本一致。这一结果符合卧龙沙棘风媒、异交的繁育系统特点。     

基于cpDNA和ITS序列对庭藤复合群的居群遗传学研究

采用2个叶绿体基因组片段(ndh J-trn F和trn D-trn T)和核基因组ITS序列对分布于中国东部和相邻地区的庭藤复合群(Indigofera decora complex)的24个居群进行分析,探讨该复合群的遗传多样性、单倍型分布、遗传结构和居群历史动态。cp DNA和ITS序列分析表明,庭藤复合群的单倍型多样性较高,而总体遗传多样性偏低(cp DNA片段:Hd=0.778,π=0.00123;ITS:Hd=0.909,π=0.01290);皖浙地区居群的遗传多样性相对较高,推测其可能是该复合群遗传多样性的分布中心。遗传变异主要存在于复合群居群间,种内居群间遗传分化较大(cp DNA片段:FST=0.848;ITS:FST=0.787);基因流较小(cp DNA片段:Nm=0.228;ITS:Nm=0.241),主要是地理隔离(遗传漂变)或生境片段化促进了物种形成;复合群的不同物种间存在由基因渐渗引起的单倍型共享现象,可能是种间不完全的生殖隔离所致。花木蓝的居群单独聚为一枝,且与其他种无共享单倍型,因此应保留花木蓝种的分类地位,而其他种的分类地位有待进一步研究。     

内蒙古高原小叶锦鸡儿(Caragana microphylla)、中间锦鸡儿(C. davazamcii)和柠条锦鸡儿(C. korshinskii)的遗传多样性及遗传关系

采用RAPD技术对10个具较大地理跨度的小叶锦鸡儿、中间锦鸡儿和柠条锦鸡儿种群的遗传多样性和遗传关系进行了研究.共检测到678个位点,多态条带比率(PPB)为100%;特有位点41个,占6.05%.总体上3种锦鸡儿的遗传多样性表现出自东向西递减的趋势,分析表明其与生长地点年均气温呈显著负相关.AMOVA表明:3种锦鸡儿种间变异只占总体变异的6.08%,且显著性检验表明这种变异不显著;种内种群间的变异占总变异的11.90%;总变异的主要部分来自种群内部(82.02%). 3种锦鸡儿各种群总体分析结果表明:种群内变异比率Hpop/Hsp为0.8013,基因分化系数Gst为0.1603,种群每代迁移数Nm为2.6192,显示种群间存在一定强度的基因流,3种锦鸡儿间表现为异交性.3种锦鸡儿多样性高低及种群聚类分布格局都表现出一定的地理连续性.     

Genetic diversity and genetic relationship of Caragana microphylla, Caragana davazamcii and Caragana korshinskii on the Inner Mongolia Plateau, China

C. microphylla, C. davazamcii and C. korshinskii exhibit a geographical replacement series from east to west on the Inner Mongolia Plateau. Currently, there is still a debate about the taxonomic and genetic relationship among these 3 species. We studied the genetic diversity and genetic relationship among these 3 species by analyzing DNA samples of individual plants from within 10 populations with random amplified polymorphic DNA (RAPD) markers. We identified 678 RAPD loci in total, of which all were polymorphic (PPB = 100%). There were 41 unique loci (6.05%). In general, a trend presented that the genetic diversity of these species decreased from east to west. Further, the genetic diversity was significantly negatively correlated with the local annual mean temperature. Analysis of molecular variation (AMOVA) showed that the genetic variation among these 3 species was only 6.08% of the total genetic variation. Between the species, the genetic variation was insignificant (P = 0.9961). The proportion of genetic variation among populations within each species was 11.90% (P < 0.001) of the total genetic variation, and the total genetic variation mainly existed within the populations (82.02%). Estimated with Shannon's index, genetic differentiation within the populations (Hpop/Hsp) was 0.8013, the coefficient of gene differentiation (Gst) was 0.1603, and the gene flow index (Nm) was 2.6192. This, thus, indicates that there is relatively high gene flow among these populations, and that these 3 species are crossbreeding. The genetic diversity level and the population distribution pattern showed geographic continuity to some extent.     

湖南茶叶植物资源的评价

对城步峒茶、江华苦茶、汝城白毛茶和云台山种等湖南茶叶植物资源的主要化学成分、光合特性和生产性状进行了研究。结果表明,茶多酚与儿茶素含量从高到低依次为汝城白毛茶、江华苦茶、城步峒茶、云台山种;各资源的游离氨基酸、茶氨酸和咖啡碱含量均为正常水平;汝城白毛茶属高茶多酚资源,它所含生物碱的组成方式与毛叶茶龙门模式种不同,虽可可碱和茶叶碱含量比茶种高,但仍以咖啡碱为主。在阴天阴凉条件下的净光合速率为汝城白毛茶＞城步峒茶＞江华苦茶＞云台山种,净光合速率与光量子通量、叶面温度呈极显著正相关,与气孔阻力呈极显著负相关,相关系数分别为０９２７８、０９１１５和－０８９３７。在晴天高温度强光条件下,净光合速率为云台山种＞城步峒茶＞江华苦茶＞汝城白毛花,净光合速率与光量子通量、叶面温度和气孔阻力都呈极显著负相关,相关系数分别为－０９１６５、－０９３２８、－０９０３１。在主要生产性状中,汝城白毛茶能安全通过－６℃的低温,在高茶多酚资源中,抗寒性突出,是选育红茶良种的一个非常有益的关键性状。     

Population genetic structure of the medicinal plant Vitex rotundifolia in China: implications for its use and conservation

Vitexrotundifolia L.is an important plant species used in traditional Chinese medicine.For its efficient use and conservation,genetic diversity and clonal variation of V.rotundifolia populations in China were investigated using inter-simple sequence repeat markers.Fourteen natural populations were included to estimate genetic diversity,and a large population with 135 individuals was used to analyze clonal variation and fine-scale spatial genetic structure.The overall genetic diversity (GD) of V.rotundifolia populations in China was moderate (GD=0.190),with about 40% within-population variation.Across all populations surveyed,the average within-population diversity was moderate (P = 22.6%; GD = 0.086).A relatively high genetic differentiation (Gst=0.587)among populations was detected based on the analysis of molecular variance data.Such characteristics of V.rotundifolia are likely attributed to its sexual/asexual reproduction and limited gene flow.The genotypic diversity (D=0.992) was greater than the average values of a clonal plant,indicating its significant reproduction through seedlings.Spatial autocorrelation analysis showed a clear within-population structure with gene clusters of approximately 20 m.Genetic diversity patterns of V.rotundifolia in China provide a useful guide for its efficient use and conservation by selecting particular populations displaying greater variation that may contain required medicinal compounds,and by sampling individuals in a population at >20 m spatial intervals to avoid collecting individuals with identical or similar genotypes.     

猕猴桃属植物的cpSSR遗传多样性及其同域分布物种的杂交渐渗与同塑

同域分布的近缘物种常常发生杂交而导致种间基因渐渗, 从而对相关物种的自然居群遗传结构产生重要影响, 近缘种间的杂交渐渗已成为进化生物学和保护生物学关注的热点。本研究采用8对cpSSR引物对我国西部高原台地向中东部丘陵平原过渡地带同域重叠分布的猕猴桃属(Actinidia)7个物种的自然居群遗传多样性、居群遗传结构和同域分布种间遗传分化进行了检测。结果表明: (1)在6个多态性位点检测到18个等位基因形成的42个单倍型, 尽管各单倍型间显示了复杂的网状进化关系, 但还是具有明显的物种特异性; (2)各物种有丰富的cpSSR遗传多样性, 但种间存在较大差异, 绵毛猕猴桃(Actinidia fulvicoma var. lanata)的遗传多样性水平最高(P = 62.50%, h_T = 0.173, H_T = 0.897), 美味猕猴桃(A. deliciosa)的最低(P = 37.5％, h_T = 0.041, H_T = 0.516); (3)尽管不同物种的居群分化程度存在较大差异, 但种内居群间存在明显分化(G_ST为0.319–0.780, F_ST为0.401–0.695), 居群间的基因流不足(N_m为0.219–0.747<1); 其中以美味猕猴桃的居群遗传分化度最高(G_ST = 0.780, F_ST = 0.695); (4)遗传分化系数G_ST(unordered alleles)与N_ST(ordered alleles)无显著差异, 揭示本研究的大多数猕猴桃属物种不存在系统地理结构, 与用Mantel检验得出的居群遗传距离和地理距离不存在显著性相关的结果一致; (5)除了中华/美味猕猴桃复合体(A. chinensis / A. deliciosa complex)的湖北五峰(HW)和广西资源(GZ)两个同域复合居群外, 同域分布的物种间遗传分化强烈(F_ST为0.476–0.990), 与UPGMA聚类时多数居群按各自物种聚类的结果一致。进一步分析表明, 中华/美味猕猴桃复合体近缘种间存在明显的共祖多态性和杂交渐渗现象, 近缘种植株分布的交错程度以及是否存在亚居群结构对杂交渐渗存在着重要影响。亲缘关系较远的物种间杂交渐渗事件稀少, 但存在个别同塑事件。本研究结果有助于进一步了解猕猴桃属植物自然居群cpDNA的遗传特性和渐渗杂交进化模式, 为我国猕猴桃野生种质资源保育及可持续开发利用提供基础数据和科学依据。