9种植物粗提物防治落叶松毛虫的试验

从生物防治的角度出发,以黑龙江省有毒杀虫植物为研究对象,从中选取9种省内分布范围较广的杀虫植物,根据其理化性质,采用索氏方法提取杀虫生理活性物质。在内蒙古赤峰市克什克腾旗经棚林场落叶松林内对落叶松毛虫(Dendrolimus superans Butler)进行了毒力测定和林间防治试验。应用兴安白头翁、朝鲜白芷、白藓、白屈菜、兴安藜芦及兴安白芷6种杀虫植物提取液稀释20倍液时,对落叶松毛虫3龄幼虫毒力作用效果较好,校正死亡率分别为85.0%、81.7%、80.0%、76.7%、73.3%和60.0%;应用兴安白头翁、白屈菜、兴安藜芦、白藓、朝鲜白芷5种杀虫植物粗提液稀释50倍时,对落叶松毛虫的防治效果分别为62.0％、60.4％、58.6％、58.6％、53.5％。并与生物药剂、植物源杀虫剂及化学农药进行了药效比较分析,其研究结果为林木害虫的生物防治及有毒植物资源的开发和利用提供理论依据。     

中国石蒜属种间亲缘关系ITS序列分析

本文利用核糖体DNA内转录间隔区(ITS)序列对石蒜属13个种(含变种)的亲缘关系进行分析。结果表明,各样品的ITS1长度为259~260 bp,ITS2为230 bp,分别有多个特异性信息位点。以ITS序列为依据对石蒜属植物亲缘关系进行分析,表明石蒜属13个种可分为三大类,其中类Ⅰ包括中国石蒜、地笑、安徽石蒜和长筒石蒜,核型为M+T型;类Ⅱ包括矮小石蒜、换锦花、玫瑰石蒜和红蓝石蒜,核型为ST型;类Ⅲ包括稻草石蒜、乳白石蒜、短蕊石蒜和两种人工杂交种,核型为ST+M+T。系统进化树与核型分析结果相似,第Ⅲ类可能为自然杂交种。     

甘草属种间杂交种叶绿体DNA父系遗传的发现及分析

通过对甘草属乌拉尔甘草(Glycyrrhiza uralensis)、光果甘草(G.glabra)、胀果甘草(G.inflata)及其人工杂交种组合G.uralensis♀×G.glabra♂、G.glabra♀×G.uralensis♂、G.uralensis♀×G.inflata♂、G.inflata♀×G.uralensis♂共68份材料的核基因ITS序列、叶绿体rbc L、mat K、trn H-psb A基因的序列分析,探讨了甘草属叶绿体DNA遗传方式。结果表明:(1)亲本种和人工杂交种ITS序列长度均为614 bp,其中34份人工杂交种ITS序列存在4处变异位点,且人工杂交种均检测出来自父本、母本ITS序列相同位点碱基的叠加,检测率为100%。(2)亲本种与人工杂交种的叶绿体基因rbc L、mat K、trn H-psb A序列长度相同,共有4处变异位点,人工杂交种在变异位点处的碱基与其相对应的父本碱基一致率高达97.1%。以上结果说明,该研究获得34份人工杂交种为100%杂交成功的F_1子代,核基因ITS序列可用于甘草属杂交种的遗传鉴定;甘草属叶绿体rbcL、mat K、trn H-psb A基因具有父系遗传特性,推测甘草属质体的遗传方式主要表现为父系遗传,这种质体遗传方式的发现为甘草属杂交种和遗传多样性研究提供了新的认识,也为杂交种的亲本鉴定提供分子依据。     

本地种云南柳与外来种旱柳（杨柳科）的同倍体自然杂交?

对分布于云南的旱柳( Salix matsudana)和云南柳( Salix cavaleriei)之间的一个自然杂交种进行了研究。野外观察表明疑似杂交种异蕊柳( Salix × heteromera)形态上介于旱柳和云南柳之间,并得到了基于叶形态特征的主成份分析的印证。核基因ITS序列数据表明这三个种存在ITS序列的种内和个体内的多态性,且疑似杂交种的ITS序列的基因型总是疑似亲本的嵌合体。因此可以判定异蕊柳是旱柳和云南柳的自然杂交后代。流式细胞分析表明这三个种均为四倍体,因而,本杂交事件为同倍体杂交。基于四个叶绿体序列片段的数据表明本自然杂交事件是不对称的,云南柳是异蕊柳的母本。常见外来种旱柳与稀有本地种云南柳的杂交可能导致稀有种云南柳的濒危甚至灭绝。研究表明柳属植物的引种应非常谨慎。     

本地种云南柳与外来种旱柳(杨柳科)的同倍体自然杂交(英文)

对分布于云南的旱柳(Salix matsudana)和云南柳(Salix cavaleriei)之间的一个自然杂交种进行了研究。野外观察表明疑似杂交种异蕊柳(Salix×heteromera)形态上介于旱柳和云南柳之间,并得到了基于叶形态特征的主成份分析的印证。核基因ITS序列数据表明这三个种存在ITS序列的种内和个体内的多态性,且疑似杂交种的ITS序列的基因型总是疑似亲本的嵌合体。因此可以判定异蕊柳是旱柳和云南柳的自然杂交后代。流式细胞分析表明这三个种均为四倍体,因而,本杂交事件为同倍体杂交。基于四个叶绿体序列片段的数据表明本自然杂交事件是不对称的,云南柳是异蕊柳的母本。常见外来种旱柳与稀有本地种云南柳的杂交可能导致稀有种云南柳的濒危甚至灭绝。研究表明柳属植物的引种应非常谨慎。     

朝鲜白头翁分生结节的诱导与组织解剖学观察

以朝鲜白头翁(Pulsatilla koreana)的叶柄为材料诱导分生结节, 利用常规石蜡切片法和树脂包埋切片法研究了朝鲜白头翁离体形态发生的解剖学特点。结果表明: 叶柄在MS+3 mg⋅L^–1玉米素(zeatin, ZEA)+0.5 mg⋅L^–1吲哚乙酸(IAA)的诱导培养基上培养42天后, 分生结节诱导率为82.5%; 培养56天后, 95%的分生结节转化成不定芽。组织解剖结构观察结果显示, 培养7、14、28和42天时分别观察到前分生结节结构、形成层和拟分生组织结构、顶端分生组织、叶原基结构。朝鲜白头翁的不定芽起源于分生结节内部的维管中心。新分化芽基部类似愈伤组织的细胞团的显微切片结果显示, 基部组织上分布着表皮细胞或下表皮细胞、维管束形成层和拟分生组织细胞, 它们在朝鲜白头翁器官分化中发挥重要作用。     

栽培种甘薯及其近缘野生种nrDNA ITS序列分析

采用核糖体DNA内转录间隔区(nrDNA ITS)序列比较分析了甘薯及其近缘野生种的遗传多样性及系统进化关系,首次报道了栽培种甘薯‘徐薯18’(Ipomoea batatas‘Xushu18’)及其近缘野生种I.triloba(DOM),I.cordatotriloba(MEX),I.nil(PER),I.nil(JPN),I.hederacea Jacq.(USA),I.hederacea Jacq.(HK)和种间杂交种67-1(I.batatas‘Xushu18’×I.hederacea Jacq.)及回交种(67-1×I.batatas‘Xushu18’)的nrDNA ITS序列。序列分析表明,栽培种甘薯及其近缘野生种nrDNA ITS序列长度为570~600bp。其中,ITS1序列为185~209 bp,GC含量为53.11%~61.83%;ITS2序列为214~226 bp,GC含量为61.21%~72.89%;5.8S序列均为165 bp,GC含量为54.55%~55.76%。此外,栽培种甘薯及其近缘野生种ITS序列信息位点均集中在ITS1和ITS2区;与其他甘薯属植物相比,I.wrightii ITS2的末端缺失了6~8个碱基。系统进化分析表明,栽培种甘薯‘徐薯18’(I.batatas‘Xushu18’)和野生种I.triloba、I.cordatotriloba、I.lacunosa、I.trifida的亲缘关系较近,与I.wrightii、I.pes-tigridis、I.grandifolia、I.nil、I.hederacea Jacq.、I.purpurea的亲缘关系较远;杂交后代与栽培种甘薯‘徐薯18’(I.batatas‘Xushu18’)亲缘关系较近,与野生种父本I.hederacea Jacq.的亲缘关系较远。     

大豆疫霉和苜蓿疫霉rDNA ITS序列分析

采用真菌核糖体基因转录间隔区(ITS)通用引物,PCR分别扩增了Phytophthora sojae的5个菌株(Pg1、Pg2、Pg3、CN1和S317)和1个P. medicaginis (菌株44390)的ITS1与ITS2,并对PCR产物进行了序列测定。根据Bioedit软件中的neighbour-joining methods分析法将上述序列和Genbank中已登录的P. sojae、P. medicaginis、P. megasperma和P.trifolii等4个形态学种10个登录菌株的ITS1与ITS2碱基序列进行聚类分析。结果是聚类组与形态学种有一定差别,4个种16个菌株分成7个聚类组。结果表明,分别属于同一形态学种且可聚为一组的不同个体之间的ITS碱基序列遗传相似性最高,但是也具有一定的多样性;形态学上属于不同种的个体的ITS可以聚为一组。上述结果提示L41385可能不属于P. sojae, L41380可能属于是P. trifolii,P. megasperma仍是一个复合种。同时提示ITS DNA碱基序列可以区分形态学种。     

大豆疫霉和苜蓿疫霉rDNA ITS序列分析*

采用真菌核糖体基因转录间隔区(ITS)通用引物,PCR分别扩增了Phytophthora sojae的5个菌株(Pg1、Pg2、Pg3、CN1和S317)和1个P. medicaginis (菌株44390)的ITS1与ITS2,并对PCR产物进行了序列测定。根据Bioedit软件中的neighbour-joining methods分析法将上述序列和Genbank中已登录的P. sojae、P. medicaginis、P. megasperma和P.trifolii等4个形态学种10个登录菌株的ITS1与ITS2碱基序列进行聚类分析。结果是聚类组与形态学种有一定差别,4个种16个菌株分成7个聚类组。结果表明,分别属于同一形态学种且可聚为一组的不同个体之间的ITS碱基序列遗传相似性最高,但是也具有一定的多样性;形态学上属于不同种的个体的ITS可以聚为一组。上述结果提示L41385可能不属于P. sojae, L41380可能属于是P. trifolii,P. megasperma仍是一个复合种。同时提示ITS DNA碱基序列可以区分形态学种。     

大豆疫霉和苜蓿疫霉rDNA ITS序列分析

采用真菌核糖体基因转录间隔区(ITS)通用引物,PCR分别扩增了Phytophthora sojae的5个菌株(Pg1、Pg2、Pg3、CN1和S317)和1个P.medicaginis(菌株44390)的ITS1与ITS2,并对PCR产物进行了序列测定。根据Bioedit软件中的neighbour-joining methods分析法将上述序列和Genbank中已登录的P.sojae、P.medicaginis、P.megasperma和P．trifolii等4个形态学种10个登录菌株的ITS1与ITS2碱基序列进行聚类分析。结果是聚类组与形态学种有一定差别,4个种16个菌株分成7个聚类组。结果表明,分别属于同一形态学种且可聚为一组的不同个体之间的ITS碱基序列遗传相似性最高,但是也具有一定的多样性;形态学上属于不同种的个体的ITS可以聚为一组。上述结果提示L41385可能不属于P.sojae,L41380可能属于是P.trifolii,P.megasperma仍是一个复合种。同时提示ITS DNA碱基序列可以区分形态学种。     

Molecular phylogeny and evolution of Caricaceae based on rDNA internal transcribed spacers and chloroplast sequence data

This study focused on clarifying phylogenetic relationships and evolution within Caricaceae. Our phylogenetic analysis based on nucleotide sequences from the ITS of the ribosomal DNA and three chloroplast fragments (matK, trnL-trnF, and psbA-trnH) included 29 taxa belonging to five genera: the neotropical genera Carica, Vasconcellea, Jarilla, and Jacaratia and the equatorial African genus Cylicomorpha. Having a relatively low mutation rate, matK, and trnL-trnF were used for estimating relationships at the generic level, while intrageneric evolution within Vasconcellea was studied with the more variable ITS and psbA-trnH sequences. Gaps, coded as binary characters, were added to the sequence alignments before performing Maximum Parsimony and Maximum Likelihood analyses. Monophyly of Caricaceae as well as phylogenetic distance between Carica and Vasconcellea species, previously belonging to the same genus, and monophyly of the resurrected genus Vasconcellea were emphasized. Within Vasconcellea, the largest genus of this family, two well-confirmed evolutionary lineages could be discerned: (1) V.xheilbornii, V. weberbaueri, V. stipulata, and V. parviflora and (2) a clade holding all other taxa of the genus. Incongruence between nuclear ITS and chloroplast psbA-trnH datasets, shown to be significantly caused by some taxa of the genus Vasconcellea, indicated that reticulate events in this genus might be more frequent than previously suspected. Moreover, intra-individual ITS sequence heterogeneity provided further evidence for the hybrid or introgressed origin of different taxa and one presumed hybrid belonging to this genus.     

Chloroplast inheritance patterns in Actinidia hybrids determined by single stranded conformation polymorphism analysis

The inheritance patterns of the chloroplast genomes of the Actinidia hybrids A. eriantha (male parent) x A. chinensis (female parent) and A. chinensis (male parent) x A. melanandra (female parent) were analyzed using single-strand conformation polymorphism (SSCP) analysis of the trnL-trnF and psbA-trnH intergenic spacers. This showed that the artificial hybrids between A. eriantha and A. chinensis all had the haplotype of their male parent. Alignment of the sequences of A. eriantha and A. chinensis revealed four substitutions and one insertion (GATTC) in trnL-trnF and two substitutions in psbA-trnH. In contrast, the haplotypes of the artificial hybrids between A. chinensis and A. melanandra had the same patterns as their female parent. Alignment of the entire region of A. chinensis and A. melanandra revealed 12 substitutions: 1 in trnL-trnF and 11 in psbA-trnH. However, no sequence variation in the trnL-trnF and psbA-trnH intergenic spacers was found. We have developed a simple screening method for detecting the inheritance patterns of Actinidia chloroplast DNA haplotypes using SSCP analysis of the trnL-trnF and psbA-trnH intergenic spacers. Our findings indicate that the inheritance of the chloroplast genome in Actinidia hybrids differs according to the species selected.     

基于叶绿体基因rbcL和psbA-trnH间区探讨石杉科植物系统关系

关于石杉科Huperziaceae植物的分类,一直存在一些争议。在旧的分类体系中石杉科植物被包含在一个混合的石松科Lycopodiaceae和多谱系的石松属Lycopodium中。本文利用叶绿体rbcL基因和psbA-trnH基因间区序列探讨石杉科植物的系统位置及石杉科内部的分类关系,用最大简约法和邻接法对自测序列结合由GenBank下载的rbcL及psbA-trnH基因间区序列进行系统发育分析。结果显示,石杉科与Phylloglossum属关系较近,与石松科关系较疏远。在石杉科中热带石杉属Huperzia植物和马尾杉属Phlegmariurus植物的关系要比它们与其他石杉属植物更近。所以,我们的rbcL基因数据不支持秦仁昌关于石杉科分为石杉属和马尾杉属的分类处理。但是,因为我们的psbA-trnH序列没有包括热带种类,对石杉属植物和马尾杉属植物的关系无验证。因此需要更多的样品和序列数据进一步探讨石杉科的演化关系。     

Genetic variation of Kaempferia (Zingiberaceae) in Thailand based on chloroplast DNA (psbA-trnH and petA-psbJ) sequences

Genetic variation and species authentication of 71 Kaempferia accessions (representing 15 recognized, six new, and four unidentified species) found indigenously in Thailand were examined by determining chloroplast psbA-trnH and partial petA-psbJ spacer sequences. Ten closely related species (Boesenbergia rotunda, Gagnepainia godefroyi, G. thoreliana, Globba substrigosa, Smithatris myanmarensis, S. supraneanae, Scaphochlamys biloba, S. minutiflora, S. rubescens, and Stahlianthus sp) were also included. After sequence alignments, 1010 and 865 bp in length were obtained for the respective chloroplast DNA sequences. Intraspecific sequence variation was not observed in Kaempferia candida, K. angustifolia, K. laotica, K. galanga, K. pardi sp nov., K. bambusetorum sp nov., K. albomaculata sp nov., K. minuta sp nov., Kaempferia sp nov. 1, and G. thoreliana, for which more than one specimen was available. In contrast, intraspecific sequence polymorphisms were observed in various populations of K. fallax, K. filifolia, K. elegans, K. pulchra, K. rotunda, K. marginata, K. parviflora, K. larsenii, K. roscoeana, K. siamensis, and G. godefroyi. A strict consensus tree based on combined psbA-trnH and partial petA-psbJ sequences revealed four major groups of Kaempferia species. We suggest that the genus Kaempferia is a polyphyletic group, as K. candida was distantly related and did not group with other Kaempferia species. Polymorphic sites and indels of psbA-trnH and petA-psbJ can be used as DNA barcodes for species diagnosis of most Kaempferia and outgroup species. Nuclear DNA polymorphism should be examined to determine if there has been interspecific hybridization and chloroplast DNA introgression in these taxa.     

秦巴山区漆树nrDNAITS和cpDNA序列分子进化特点的分析

应用PCR产物直接测序法分析漆树种群nrDNA （核糖体DNA） ITS序列和cpDNA序列（matK、rbcL、psbA-trnH）的碱基差异,初步研究两套植物基因组的变异速率。结果表明：nrDNA ITS序列共有518 bp,有变异位点15处,变异位点百分率为2.9%,（G+C）含量为61.8%。cpDNA序列合并后长度1 907 bp,有变异位点20处,变异位点百分率为1.05%,（G+C）含量为36.1%。通过对ITS序列核糖型（Ribotype）和叶绿体序列单倍型（Haploype）进行分析发现,秦巴山区漆树区域性分布明显,不同区域拥有自己独特的单倍型,漆树居群历史近期没有扩张。nrDNA ITS序列较叶绿体序列进化较快,变异速率较快。nrDNA ITS序列及叶绿体matK、psbA-trnH适合漆树的亲缘地理学研究。     

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.     

鸡爪槭种下分类群的DNA条形码筛选

槭属鸡爪槭(Acer palmatum Thunb.)是北温带广泛分布的一类园艺观赏树种,由于频繁的种内杂交渐渗导致其种下分类群的形态性状特征趋同,致使传统的形态学分类难以准确鉴定,新兴的DNA条形码技术为快速、准确的鉴定鸡爪槭种下分类群提供了新的思路。本研究采用5个叶绿体DNA片段(rpl16、psbA-trnH、trnL-trnF、rbcL、matK)和核基因组ITS片段,运用PWG-distance和Tree-Building两种方法对鸡爪槭的8个分类群共32个个体进行DNA条形码分析。结果显示,单个叶绿体基因组片段(分辨率为0%~25%)或核rDNA ITS片段(12.5%)的分辨率较低,不同组合的叶绿体DNA片段(0%~62.5%)、叶绿体片段与核rDNA ITS片段(12.5%~50%)的分辨率则相对较高。其中,rpl16+psbA-trnH+trnL-trnF片段组合的分辨率最高(62.5%),较为符合DNA条形码快速、准确鉴定的要求,因此建议将其作为鸡爪槭种下分类群鉴定的DNA条形码。     

Sequence characteristics and divergent evolution of the chloroplastpsbA-trnH noncoding region in gymnosperms

ThepsbA-trnH intergenic region is among the most variable regions in the gymnosperm chloroplast genome. It is proposed as suitable for DNA barcoding studies and is useful in phylogenetics at the species level. This region consists of two parts differing in their evolutionary characteristics: 1) thepsbA 3′UTR (untranslated region) and 2) thepsbA-trnH intergenic spacer. We compared the sequence and RNA secondary structure of thepsbA 3′ UTR across gymnosperms and found consensus motifs corresponding to the stem portions of the RNA stem-loop structures and a consensus TGGATTGTTATGT box. ThepsbA-trnH spacer is highly variable in length and composition. Tandem repeats that form stem—loop structures were detected in both thepsbA 3′ UTR and the psbA-trnH spacer. The presence of promoters and stem—loop structures in the psbA-trnH spacer and high sequence variation in this region suggest that psbA and trnH in some gymnosperms are independently transcribed. Acomparison of chloroplast UTRs across gymnosperms offer clues to the identity of putative regulatory elements and information on selective constraints imposed on the chloroplast non-coding regions. The present study should inspire researchers to explore the full potential of thepsbA-trnH non-coding sequence and to further stimulate its application in a broader spectrum of studies, not limited to phylogenetics and DNA barcoding.     

Molecular evidence for the maternal parentage in the hybrid origin of Spartina anglica C.E. Hubbard

Spartina anglica is a textbook example of a natural amphiploid, which originated from hybridization between S. alterniflora and S. maritima . Which of these species was the maternal parent has remained a mystery. Inheritance of chloroplast DNA in most angiosperms is strictly maternal and can thus be used to test the parentage of hybrid taxa. The DNA sequence of the chloroplast leucine tRNA gene intron was used to show that the introduced North American S. alterniflora is the female parent of the F ₁ hybrid S. x townsendii and the amphiploid S. anglica . A possible scenario for their origin is given.     

Recombination and introgression of nuclear and chloroplast genomes between the peat mosses, Sphagnum capillifolium and Sphagnum quinquefarium

Haploid hybrid gametophytes are often present at low frequencies in sympatric populations of Sphagnum capillifolium and Sphagnum quinquefarium. We used intersimple sequence repeat (ISSR) markers and polymerase chain reaction-restriction fragment length polymorphism of the trnL(UAA) intron of the chloroplast genome to reveal the nuclear and chloroplast composition of mature hybrid gametophytes from natural populations and of gametophytes derived from spores of hybrid sporophytes collected in nature. Asymmetrical nuclear inheritance was found in the progeny of the hybrid sporophytes, indicating that only spores with a low level of recombination of parental genomes were viable. A similarly skewed nuclear composition was found among the naturally occurring hybrid gametophytes. All hybrid genomes contained a larger proportion of S. capillifolium ISSR markers, combined with only two to five S. quinquefarium markers together with a chloroplast haplotype derived from S. quinquefarium. In this way, a pattern resembling introgression is created within a single generation. Some individuals possessed nuclear genomes typical for S. capillifolium in combination with the chloroplast haplotype of S. quinquefarium, possibly indicating backcrossing. Our results indicate that hybridization between S. capillifolium and S. quinquefarium is relatively common, but the resistance of large parts of the genome against heterospecific genes maintains the genetic distinctness of the species. Further evolutionary and phylogenetic consequences of restricted interspecific gene exchange are discussed.