中国鸟类的DNA分类及系统发育研究概述

鸟类分类是鸟类学其他研究领域的基础,近年来分子技术的发展,以及计算机技术的应用为鸟类分类学和鸟类系统演化研究提供了新的研究手段,给传统的系统分类研究带来了新的机遇.Tautz等于2002年首先提出运用DNA序列作为生物分类系统的主要平台,即DNA分类学(DNA Taxonomy).而Hebert等于2003年则首次提出了DNA条形码(DNA Barcoding)的概念,并对其物种分类和鉴定意义予以肯定,建议利用线粒体细胞色素C氧化酶亚单位Ⅰ(COI)的特定区段来做DNA条形编码的基础.在鸟类DNA分类方面,国内学者应用线粒体基因Cut b,COI,c-mos,c-myc,12s rRNA,16s rRNA,ND2,ND3,CR,RAG-1以及核基因myoglobin introⅡ等不同片段对很多类群进行了分类探讨和系统发育研究.但是主要集中在鸡形目及雀形目鸟类.中国是鸟类多样性极其丰富的国家,近年来很多亚种、种及以上分类阶元依然存在问题,因此,中国鸟类物种的分类地位、系统发育与演化关系等依然有很多问题等待深入研究.目前国内基于COI的鸟类分类及系统发育研究有了一些报道,但是真正的DNA条形码工作尚需继续、深入地开展.

Phylogeny of east Asian bufonids inferred from mitochondrial DNA sequences (Anura : Amphibia)

We investigated the relationships of Asian bufonids using partial sequences of mitochondrial DNA genes. Twenty-six samples representing 14 species of Bufo from China and Vietnam and 2 species of Torrentophryne from China were examined. Three samples of Bufo viridis from Armenia and Georgia were also sequenced to make a comparison to its sibling tetraploid species B. danatensis. Bufo americanus, from Canada, was used as the outgroup. Sequences from the 12S ribosomal RNA, 16S ribosomal RNA, cytochrome b, and the control region were analyzed using parsimony. East Asian bufonids were grouped into two major clades. One clade included B. andrewsi, B. bankorensis, B. gargarizans, B. tibetanus, B. tuberculatus, its sister clade B. cryptotympanicus, and the 2 species of Torrentophryne. The second clade consisted of B. galeatus, B. himalayanus, B. melanostictus, and a new species from Vietnam. The placement of three taxa (B. raddei B. viridis, and its sister species, B. danatensis) was problematic. The genus Torrentophryne should be synonymized with Bufo to remove paraphyly. Because B. raddei does not belong to the clade that includes B. viridis and B. danatensis, it was removed from the viridis species group. The species status of B bankorensis from Taiwan is evaluated. (C) 2000 Academic Press.

Meiothermus rosaceus sp nov isolated from Tengchong hot spring in Yunnan, China

A rosy-pigmented Gram-negative, thermophilic bacterium with an optimum growth temperature of about 55degreesC was isolated from Tengchong hot springs in Yunnan province, China. Its growth scarcely occurred below 40degreesC or above 70degreesC. Phylogenetic and secondary structural analyses of 16S rRNA and DNA-DNA hybridization showed that the organism represented a new species of the genus Meiothermus. This new species could be distinguished easily from other species of the genus Meiothermus by the following phenotypic characteristics: rosy pigment, expanded body, sucrose and maltose were not utilized, gelatin and starch were not hydrolyzed. On the basis of the above data, the name Meiothermus rosaceus sp. nov. was proposed for the species represented by the strain RH9901(T)(CCTCC-AB200291). (C) 2002 Federation of European Microbiological Societies. Published by Elsevier Science B.V. All rights reserved.

The Phylogenetic Relationships of the Chinese and Vietnamese Waterfall frogs of the genus Amolops

Ranid frogs of the genus Amolops occur in Southeast Asia and are typically found near waterfalls. Their phylogenetic relationships have not been resolved. We include 2,213 aligned nucleotide sites of the 12S, 16S and tRNA(val) gene regions of the mitochondrial DNA genome from 43 individuals of Chinese and Vietnamese Amotops, Huia, Hylarana, Meristogenys, Odorrana and Rana. The outgroup species were from the genera Chaparana, Limnonectes, Nanorana, and Paa. The data were analyzed within the framework of a refutationist philosophy using maximum parsimony. Four clades of waterfall frogs were resolved. Meristogenys was not resolved as the sister group to either Huia nor Amolops. The hypothesis Of evolutionary relationships placed Amolops chapaensis and Huia nasica in the genus Odorrana.

原生动物单细胞PCR应用初探——以天蓝喇叭虫为例

以天蓝喇叭虫(Stentor coeruleus)为研究对象,探索了单细胞单基因PCR扩增及单细胞全基因组PCR扩增技术在原生动物中的应用.经过不断探索和优化条件后,试验取得了理想的结果.在40例单细胞单基因(SSU rDNA基因全序列)PCR的一次性扩增中,新鲜细胞和经过中性红染色的细胞都获得了100%的成功率,室温下酒精(95%)保存一周的细胞获得了82.5%的成功率.在单细胞全基因组PCR扩增中,采用高效高保真的phi29DNA聚合酶结合随机引物(Random Primer)进行扩增,获得了丰富且质

中国9种嗜子宫线虫系统发育的初步研究

为了探讨鱼类寄生嗜子宫线虫的系统发育关系,测定了8种嗜子宫线虫的ITS rDNA(核糖体转录内间隔区核 糖核酸)序列和9种嗜子宫线虫的18S rDNA(小亚基核糖体核糖核酸)部分序列,并构建了18S rDNA序列的系统发 育树。在比较和分析ITS rDNA和18S rDNA两种分子标记对嗜子宫科线虫系统发育适用性的基础上,分析了嗜子 宫线虫的系统发育关系。结果表明:中国嗜子宫线虫是单系起源;黄颡鱼似嗜子宫线虫、赣州似嗜子宫线虫和棍头 嗜子宫线虫亲缘关系非常接近,可能是较晚形成的种;似嗜子宫线虫属可能应该被

全细胞PCR扩增用于鱼腥藻种类鉴定

传统鉴定藻种的方法主要是通过形态学观察的方法加以判断。蓝藻在自然条件和人工培养过程中 ,其形态、代谢能力等都可能发生变化 ,同时该过程需要的时间长 ,难以区分种或种以下的分类、单位 ,亦难以在水华暴发早期阶段准确鉴定。本文利用rDNA通用引物扩增 ,表明在 5 0 μL的反应体系中加入 2 0个鱼腥藻细胞能扩增出目的条带 ;对已知的鱼腥藻PC基因的分析设计引物 ,在BSA浓度为 0 2 %— 1% (w/v)下 ,全细胞扩增出实验室保存的四种鱼腥藻的部分PC以及PC IGS序列 ,序列分析结果表明PC I

水华藻类的分子鉴定研究进展

中国科学院知识创新工程重大项目 (KZCX1 SW 12 ); 973项目 (2 0 0 2CB412 3 0 0 ); 国家重大环境课题“滇池蓝藻水华污染控制技术研究”(K99 0 5 3 5 0 1); 中国科学院方向性创新课题 (No .2 2 0 3 16)资助

中国石爬鮡属鱼类的形态变异及物种有效性研究

石爬属鱼类的青石爬和黄石爬的物种界限一直不清楚。采用形态判别和线粒体 16SrRNA基因序列分析相结合的方法 ,分别研究分析了青石爬和黄石爬的物种划分、地理分化及遗传多态性。结果表明 :(1)区别青石爬和黄石爬的重要特征腹鳍起点至臀鳍起点的距离是否大于至鳃孔下角的距离 ,腹鳍相对位置 ,头部相对大小与上颌须的须状延长部分等相互之间有一定的相关性 ,但是在研究的样本中没有截然的界限 ,而是有较大的重叠 ,难以区分 ;(2 )从地埋分布看 ,金沙江不同支流的样本在上述特征上有一定的区别 ,但是没

四种臂尾轮虫rDNA16S-23S基因间隔区的序列测定与分析

中国科学院生物分类区系学科发展特别支持项目

二种淡水微囊藻rDNA16S-235基因间隔区的序列测定与分析

本研究采用ＰＣＲ及序列测定的方法，对我国淡水铜绿微囊藻有毒株（Ｍ８６４１）和另一低毒的种类惠氏微囊藻（Ｍ５７４）ｒＤＮＡ１６Ｓ－２３Ｓ基因间隔区进行了序列的测定和分析，结果表明：ｒＤＮＡ１６Ｓ－２３Ｓ基因间隔区可以作为一个精细且稳定的指标，用于微囊藻的分类和鉴定。并从分子水平提出了铜绿微囊藻与惠氏微囊藻在种系发生上有较近缘的关系。本文首次对微囊藻属Ｍｉｃｒｏｃｙｓｔｉｓ　ｒＤＮＡ基因间隔区全序列作了报道，为微囊藻属的鉴定及系统学研究提供了分子基础。

蓝藻OscilatoriaceaerDNA16S-23S基因间隔区的序列分析及其分类学意义

采用末端终止法对蓝藻类颤藻科Ｏｓｃｉｌａｔｏｒｉａｓｐ．ｒＤＮＡ１６Ｓ－２３Ｓ基因间隔区进行了序列测定，获得了Ｏｓｃｉｌａｔｏｒｉａｓｐ．ｒＤＮＡ基因间隔区４２７个核苷酸，其中包含１个异亮氨酸ｔＲＮＡ基因（ｔＲＮＡＩｌｅ）。并通过计算机联网从国际分子生物学数据弹库中获取颤藻科其它种的ｒＤＮＡ基因间隔区序列，通过比较分析，从分子水平对颤藻科Ｏｓｃｉｌａｔｏｒｉａｃｅａｅ属间的某些分类学问题进行了讨论，并根据序列中核苷酸差异值探讨了颤藻科属间界定的分子标准。提出了ｒＤＮＡ基因间隔区是良好的分子标记，可用于“赤

PCR-DGGE Fingerprinting Analysis of Plankton Communities and Its Relationship to Lake Trophic Status

Plankton communities in eight lakes of different trophic status near Yangtze, China were characterized by using denatured gradient gel electrophoresis (DGGE). Various water quality parameters were also measured at each collection site. Following extraction of DNA from plankton communities, 16S rRNA and 18S rRNA genes were amplified with specific primers for prokaryotes and eukaryotes, respectively; DNA profiles were developed by DGGE. The plankton community of each lake had its own distinct DNA profile. The total number of bands identified at 34 sampling stations ranged from 37 to 111. Both prokaryotes and eukaryotes displayed complex fingerprints composed of a large number of bands: 16 to 59 bands were obtained with the prokaryotic primer set; 21 to 52 bands for the eukaryotic primer set. The DGGE-patterns were analyzed in relation to water quality parameters by canonical correspondence analysis (CCA). Temperature, pH, alkalinity, and the concentration of COD, TP and TN were strongly correlated with the DGGE patterns. The parameters that demonstrated a strong correlation to the DGGE fingerprints of the plankton community differed among lakes, suggesting that differences in the DGGE fingerprints were due mainly to lake trophic status. Results of the present study suggest that PCR-DGGE fingerprinting is an effective and precise method of identifying changes to plankton community composition, and therefore could be a useful ecological tool for monitoring the response of aquatic ecosystems to environmental perturbations.

Identification of Prorocentrum minimum and Takayama pulchella by fluorescence in situ hybridization through epifluorescence microscopy and flow cytometry

Partial rDNA sequences of Prorocentrum minimum and Takayama pulchella were amplified, cloned and sequenced. and these sequence data were deposited in the GenBank. Eight oligonucleotide probes (DNA probes) were designed based on the sequence analysis. The probes were employed to detect and identify P. minimum and T. pulchella in unialgal and mixed algal samples with a fluorescence in situ hybridization method using flow cytometry. Epifluorescence micrographs showed that these specific probes labeled with fluorescein isothiocyanate entered the algal cells and bound to target sequences, and the fluorescence signal resulting from whole-cell hybridization varied from probe to probe. These DNA probes and the hybridization protocol we developed were specific and effective for P. minimum and T. pulchella, without any specific binding to other algal species. The hybridization efficiency of different probes specific to P. minimum was in the order: PM18S02 > PM28S02 > PM28S01 > PM18S01, and that of the probes specific to T. pulchella was TP18S02 > TP28S01 > TP28S02 > TP18S01. The different hybridization efficiency of the DNA probes could also be shown in the fluorescent signals between the labeled and unlabeled cells demonstrated using flow cytometry. The DNA probes PM18S02, PM28S02; TP18S02 and TP28S01, and the protocol, were also useful for the detection of algae in natural samples.

Morphological and molecular phylogenetic analysis of two Saprolegnia sp (Oomycetes) isolated from silver crucian carp and zebra fish

Two Saprolegnia isolates, JY isolated from silver crucian carp (Carassius auratus gibelio Bloch) and BMY isolated from zebra fish (Brachydanio rerio Hamilton) came from infections occurring concurrently in different locations in China. To confirm whether the two isolates were from the same Saprolegnia clone, comparative studies have been carried out based on their morphological, physiological and molecular characteristics. Observations showed that morphologically (both asexual and sexual organs) the two isolates were broadly similar and both isolates under-went repeated zoospore emergence. Comparing 704 base pairs of internal transcribed spacer (ITS) region and the 5.8S rDNA, we found isolates JY and BMY shared an identical ITS sequence with a minor variation (99.6 % similarity). Forty available sequences for representatives Saprolegnia spp. belonged to four phylogenetically separate clades. The two studied isolates fell within clade I that comprised a group of isolates which showed almost an identical ITS sequence but had been identified as a number of different morphological species. our findings suggest that isolates JY and BMY appear to belong to the S. ferax clade and this clade (1) contains a number of closely related phylogenetic species. This is distinct from the more common fish pathogenic isolates, which belong to the S. parasitica clade (III) and are characterized by having cysts decorated by bundles of long hooked hairs and two further clades (II and IV) containing largely saprotrophic or soil born species. (C) 2009 Published by Elsevier Ltd on behalf of The British Mycological Society.