Molecular differentiation of the microgastrine species commonly found in paddy fields from Southeast Asia,with additional data on their phylogeny （Hymenoptera：Braconidae）

Partial DNA sequences of three genes, that is, mitochondrial large ribosomal subunit (16S), nuclear large ribosomal subunit (28S D2) and mitochondrial NADH1 dehydrogenase (NADH1) gene, were sequenced from different microgas trine species(Braconidae: Microgastrinae) collected fresh from paddy fields. The DNA sequences were used to determine the extent of sequence variation among species in order to evaluate the specific status of each species. Cladistic analysis was also used to infer a phylogenetic relationship among these species. The results showed that sequence divergence among species of the same genus Cotesia was much lower than those among different genera, such as Cotesia, Exoryza and Apanteles; the sequence similarity of 16S rDNA and NADH 1 genes between Cotesia sp. and C. chilonis was higher than that between C. sp. and C. ruficrus.Phylogenetic analyses suggested that four species of Cotesia were always grouped in the same clade regardless of using different analysis methods; Cotesia sp. and C. chilonis are more closely related to each other than to C. ruficrus, different from previous morphological results. Additionally, sequence analyses indicated that NADH1 gene has more parsimony informative characters than 28S rDNA D2 and 16S rDNA at the species-level analysis,indicating that NADH1 gene might be a useful marker for species-level analysis.     

Comparison of Skull Morphology in Two Species of Genus Liua(Amphibia:Urodela: Hynobiidae),L.shihi and L.tsinpaensis

Skull characteristics play an important role in the systematics of tailed salamanders. In this study, the skulls of Liua shihi and L.tsinpaensis were compared using a clearing and double-staining technique. The results showed that in L.tsinpaensis, the vomerine tooth rows are in a "⌒⌒" shape, the length of the inner vomerine tooth series is nearly equal to that of the outer series, the vomerine tooth rows do not extend beyond the choanae, an ossified articular bone is absent, the basibranchial is rod shaped, the radial loops exhibit a figure-eight shape, the cornua has two cylindrical branches, the urohyal is rod shaped, and the end of the ceratohyal is not ossified; these features differ considerably from those of L.shihi. The ossification of the posterior portion of the ceratohyal and the present or absent of ossified articular might represent ecological adaptation to feeding in different environments.     

Characterization of the Agamid Lizard Genus Uromastyx from Eastern Sudan Based on Morphology,Karyotypes and Mitochondrial DNA

This study focuses on taxonomy of Uromastyx species. Morphologically, U. ocellata and U. ornata were identified and cytogenetically, 2n = 36 chromosome. 16S, cyt b and 12S genes fragmentamplification shows 597, 399 and 450 base pair respectively and more A-T than G-C base pair andmore A/C than T/G base contents. While, sequences blasting in GenBank, followed by construction of phylogenetic tree revealed, 16S and cry b sequences were similar to U. ocellata sequences, andclustered together in phylogenetic tree, in contrast, 12S sequences from same species were related tosubspecies U. ornata ornata sequences and clustered together in the phylogenetic tree. Therefore, the results of this study suggest more studies on Uromastyx species in Sudan.     

三种猕猴属动物Fas基因的克隆

从腹股沟淋巴结或血液中提取总RNA,根据人的Fas基因的cDNA序列设计上、下游引物,通过RT-PCR扩增出三种猕猴属动物Fas基因的cDNA片段,将片段克隆到pGEM-TEasy载体中,筛选阳性克隆并进行序列测定。恒河猴Fas基因编码序列为1005bp,GenBank收录号为AY007572;熊猴的编码序列为996bp,GenBank收录号为AF326208;短尾猴的编码序列为933bp,GenBank收录号为AF332357。对五种已知的灵长类动物的Fas基因进行序列比较,结果表明Fas基因的保守区域具有高度保守性,而处于编码序列中间的一段低复杂度序列在不同的物种间具有很高的变异性。 Abstract：Total RNA were extracted from inguinal lymph nodes o r blood.The Fas cDNA fragments of three macaca species were obtained by RT-P CR using the upper and low primers according to the Fas gene of human,and th en cloned into pGEM-T Easy vector.The positive clones were sequenced.For the fir st time we cloned the Fas genes of Macaca mulatta,Macaca assamensis and Macaca arctoides,the encoding sequences are 1005bp, 996bp and 933bp, respect ively.Finally we compared the sequences of Fas genes among five primate spec ies that have already been submitted to GenBank.The results confirmed the conser vation of regions in two ends of Fas genes,and the high variability of a low complexity region in middle of every Fas gene.     

Phylogenetic relationships among six species of Epistylis inferred from 18S-ITS1 sequences

Phylogenetic relationships among six species of Epistylis (i. e. E. plicatilis, E. urceolata, E. chrysemydis, E. hentscheli, E. wenrichi, and E. galea) were investigated using sequences of the first internal transcribed spacer region (ITS-1) of ribosomal DNA (rDNA). Amplified rDNA fragment sequences consisted of 215 or 217 bases of the flanking 18S and 5.8S regions, and the entire ITS-1 region (from 145 to 155 bases). There were more than 33 variable bases between E. galea and the other five species in both the 18S region and the ITS-1 region. The affiliation of them was assessed using Neighbor-joining (NJ), maximum parsimony (MP) and maximum likelihood (ML) analyses. In all the NJ, MP and ML analyses E. galea, whose macronucleic position and shape are distinctly different from those of the other five species, was probably diverged from the ancestor of Epistylis earlier than the other five species. The topology in which E. plicatilis and E. hentscheli formed a strongly supported sister clade to E. urceol     

Variations of 18S rDNA Loci Among Six Populations of Paeonia obovata Maxim. （Paeoniaceae） Revealed by Fluorescence In Situ Hybridization

The localization of 18S ribosomal RNA genes （rDNA） by fluorescence in situ hybridization （FISH） had been performed for some species of Paeonla. However, the pattern of 18S rDNA loci among populations Is Indistinct. In the present study, we localized 18S rDNA loci on meiotic or mitotic chromosomes of six populations of Paeonla obovata Maxim. （Paeonlaceae）. Different numbers of rDNA loci were found with different diploid （2n=10） populations, namely eight （Lushl and Mt. JIuhua populations）, 10 （Mt. Talbal population）, and seven （Mt. Guandl population）, whereas tetraplold （2n=20） populations were all found with 16 loci. Aii rDNA loci were mapped near teiomeres of mitotic chromosomes and there was no chromosome with two loci. The present results show that molecular cytological polymorphlsm exists among P. obovata diploid populations, Indicating that structural variations occurred frequently during the evolutionary history of this species, accompanied with differentiation among populations.     

从线粒体基因探讨中国大头蛙群的分类及其属内地位

Partial sequences of the mitochondrial 12S rRNA and 16S rRNA gene were determined for 8 populations of three species of Chinese Limnonectes, and aligned with the published sequences of Limnonectes from other parts of the world. When Nanorana parker, Paa boulengeri, Fejervarya limnocharis and Hoplobatrachus rugulosus was used as outgroup taxa (Accession Nos. AY158705, AY313685, AF206111, AF206491, AY322311). The sequences of the 12S rRNA and 16S rRNA genes totaled 950 nueleotide positions with gaps including 510 variable sites. We reconstructed phylogenetie trees using Clustal X 1.8, Mega 2.1 and PHYLIP 3.5e software, and using the maximum parsimony and maximum likelihood methods, respectively. Our analyses suggest that these fanged frogs from China are another monophyletie group in addition to the four monophyletie groups identified by previous studies. The Chinese Limnonectes were grouped into three elades (BCL 55% ). The first elade contains one species (BCL 100% ), from a population of Limnoneetes fragilis from Hainan Province. The second contains four individuals (BCL 100% ), i. e. two populations of Limnonectes kuhlii from Yunnan Province. The third contains one species (BCL 100% ), i. e. five populations of Limnonectes fujianensis from Fujian Province and 1 from Taiwan Province. The resulted phylogenetie trees indicate L. fragilis is basal to L. kuhlii L. fujianensis 。     

Molecular systematics analysis of Lymantria dispar based on 18S rRNA and cox1 mtDNA sequence data

The complete sequence of the 18 S ribosomal RNA gene(18S rRNA) from Lymantria dispar was cloned and analysed here. 18 S rRNA and mitochondrial cytochrome c oxidasel(cox1) gene sequences of Lymantria dispar were compared with homologous sequences of other nine insects from different orders. Analytic results showed that 18 S rRNA of these insects had two conserved domains and the second domain was an even more conserved region. The phylogenetic trees based on the full-length sequence and the second domain fragment of 18 S rRNA as well as sequence of cox1 from different orders indicated that Lepidoptera and Trichoptera, which belongs to Amphiesmenoptera, had a closer phylogenetic relationship and fewer differences were observed comparing with traditional taxonomic results.     

Loss of Genetic Diversity of Domesticated Panax notoginseng F H Chen as Evidenced by ITS Sequence and AFLP Polymorphism： A Comparative Study with P. stipuleanatus H T Tsai et K M Feng

In the present study, we evaluated the genetic diversity of Panax notoginseng F H Chen, a domesticated species, and P. stipuleanatus H T Tsai et K M Feng, an endangered wild species in southeastern Yunnan and adjacent areas in Vietnam, using sequences of the internal transcribed spacer (ITS) regions of nuclear ribosomal DNA and amplified fragment length polymorphism (AFLP) markers. Twenty-four accessions from three plantations of P. notoginseng and 51 samples from eight populations of P. stipuleanatus were assayed. A total of 694 bp of partial sequences of 18S, ITS 1, 5.8S, ITS2, and partial sequences of 26S were obtained. No sequence variation was detected within P. notoginseng and nine sites (1.30%) were variable in P. stipuleanatus. Two-thirds of the variable sites were found between Langqiao and other populations. In P. notoginseng, four pairs of AFLP primer combinations generated 312 bands, of which 240 (76.9%) were polymorphic and 60.15% of the polymorphisms were harbored within plantations. Approximately 41.0% and 66.9% of bands were polymorphic in population D7 and 5589, respectively. In P.stipuleanatus, the same four primer combinations produced 346 bands, of which 334 (96.5%) were polymorphic and approximately 62.14% of polymorphisms were maintained within populations. Considerable variations were observed. The percentage of polymorphic bands ranged from 50.2% to 84.9% and the average over populations was 70.9%. Cluster analysis did not show correlation of genetic differentiation with the distinctive leaf morphology of P. stipuleanatus (i.e. one form with bipinnatifid leaflets and the other with undivided leaflets). Because over 40% of genetic variations were maintained among populations and because of the very restricted distribution of P. stipuleanatus, all natural populations of this species should be conserved in situ. Considering that there are variations in P. notoginseng within and among plantations, we suggest establishing a genetic resource conservation garden or reintroducing P. notoginseng into its native habitats in southwestern China. Such reintroduction should be carefully executed after large-scale screening of genetic variation within the species.     

The first mitochondrial genome for the butterfly family Riodinidae(Abisara fylloides) and its systematic implications

The Riodinidae is one of the lepidopteran butterfly families. This study describes the complete mitochondrial genome of the butterfly species Abisara fylloides, the first mitochondrial genome of the Riodinidae family. The results show that the entire mitochondrial genome of A. fylloides is 15 301 bp in length, and contains 13 protein-coding genes, 2 ribosomal RNA genes, 22transfer RNA genes and a 423 bp A+T-rich region. The gene content, orientation and order are identical to the majority of other lepidopteran insects. Phylogenetic reconstruction was conducted using the concatenated 13 protein-coding gene(PCG) sequences of19 available butterfly species covering all the five butterfly families(Papilionidae, Nymphalidae, Peridae, Lycaenidae and Riodinidae). Both maximum likelihood and Bayesian inference analyses highly supported the monophyly of Lycaenidae+Riodinidae,which was standing as the sister of Nymphalidae. In addition, we propose that the riodinids be categorized into the family Lycaenidae as a subfamilial taxon.     

我国有尾动物分类中几个问题的研究

本文对有尾两栖类小鲵科Hynobiidae山溪鲵属、巴鲵属、小鲵属分类研究中的同物异名,以及属称的订正方面进行了进一步探讨,认为我国山溪鲵属应为5种,无唇褶山溪鲵B.cochranae应为北方山溪鲵Batrachuperus ti-betanus同物异名。巴鲵属是一个有效属称,义乌小鲵Hynobius yiwuensis应为中国小鲵H.chinensis的同物异名,黄斑拟小鲵Pseudohynobius flavonaculatus可以暂定为秦巴北鲵Ranodon tsinoaensis的同物异名。黑抓异鲵Xenobius melanonychus是否有效种有待确定。     

Heterogeneity of the conserved ribosomal ribonucleic acid sequences of Bacillus subtilis

Doi, Roy H. (University of California, Davis), and Richard T. Igarashi. Heterogeneity of the conserved ribosomal ribonucleic acid sequences of Bacillus subtilis. J. Bacteriol. 92:88-96. 1966.-Hybrid formation was demonstrated between Bacillus subtilis ribosomal ribonucleic acid (RNA) and deoxyribonucleic acid (DNA) from various bacterial species. The high degree of complementarity between B. subtilis ribosomal RNA and the DNA from B. cereus and B. stearothermophilus suggested a method to test whether the same RNA sequences were hybridizing with the DNA from these two species. Saturation studies with 16S and 23S RNA preparations from B. subtilis showed that a definite number of complementary sites was present in each DNA. Base composition analyses of the RNA in the hybrid demonstrated that ribosomal RNA sequences were involved. Hybrid competition studies revealed that B. stearothermophilus ribosomal RNA could compete totally against B. subtilis ribosomal RNA for B. stearothermophilus DNA, although it could compete only partially against the B. subtilis ribosomal RNA hybridizing with B. cereus DNA. These observations were made independently with both 16S and 23S ribosomal RNA preparations. These results revealed that different nucleotide sequences of B. subtilis ribosomal RNA were hybridizing with the DNA from B. cereus and B. stearothermophilus. Two possible interpretations of these results are: (i) different nucleotide sequences from a homogeneous ribosomal RNA population are hybridizing with heterologous DNA preparations, and (ii) ribosomal RNA cistrons are heterogeneous.     

大头蛙和脆皮大头蛙线粒体3个基因的测定及两栖类亲缘关系研究

现存的两栖类系统发生关系一直存在争议,特别是3个目间的亲缘关系。本文设计了5对引物,扩增和测定了大头蛙和脆皮大头蛙线粒体12S和16S rRNA基因和Cytb基因的全序列。在对所测序列进行分析的同时,基于3个基因全序列的相加数据,运用MEGA 3.1和PHYLIP 3.64软件中的NJ法、MP法和ML法,对两爬类17个物种,以鱼类非洲肺鱼为外群,重建出3个树形完全一致的分子系统树。研究结果显示:现存两栖类中无尾目和有尾目为姐妹群关系,并推断有尾目内小鲵科和隐鳃鲵科亲缘关系较近。此外,在研究两栖类系统发生关系方面,作者分析前人研究中产生两种不同观点的可能原因,同时总结了在此类研究中产生偏差的几种影响因素。     

Use of 16S Ribosomal DNA for Delineation of Marine Bacterioplankton Species

All of the marine bacterioplankton-derived 16S ribosomal DNA sequences previously deposited in GenBank were reanalyzed to determine the number of bacterial species in the oceanic surface waters. These sequences have been entered into the database since 1990. The rate of new additions reached a peak in 1999 and subsequently leveled off, suggesting that much of the marine microbial species richness has been sampled. When the GenBank sequences were dereplicated by using 97% similarity as a cutoff, 1,117 unique ribotypes were found. Of the unique sequences, 609 came from uncultured environmental clones and 508 came from cultured bacteria. We conclude that the apparent bacterioplankton species richness is relatively low.     

Use of 16S ribosomal DNA for delineation of marine bacterioplankton species

All of the marine bacterioplankton-derived 16S ribosomal DNA sequences previously deposited in GenBank were reanalyzed to determine the number of bacterial species in the oceanic surface waters. These sequences have been entered into the database since 1990. The rate of new additions reached a peak in 1999 and subsequently leveled off, suggesting that much of the marine microbial species richness has been sampled. When the GenBank sequences were dereplicated by using 97% similarity as a cutoff, 1,117 unique ribotypes were found. Of the unique sequences, 609 came from uncultured environmental clones and 508 came from cultured bacteria. We conclude that the apparent bacterioplankton species richness is relatively low.     

Variability and functional role of chromosomal sequences in 16SrI-B subgroup phytoplasmas including aster yellows and related strains

AIMS: Partial genetic characterization of several chromosomal regions on 35 16SrI-B phytoplasma strains maintained in periwinkle and collected in different geographical areas from plants of diverse species. METHODS AND RESULTS: Genes coding for ribosomal protein rpL22, elongation factor EF-Tu and random cloned sequences amplified with primers AY19p/m, G35p/m and BB88F1/R1 after RFLP analyses showed a high degree of polymorphism among the strains studied. The ribosomal protein (rp) subgroups B and K, and an undescribed subgroup designated N, were identified. Amplicons obtained with primers AY19p/m and BB88F1/R1, revealed a high and a low degree of polymorphism, respectively. CONCLUSIONS: A probable spacer role could be attributed to the AY19p/m sequence and a possible coding function to the BB88F1/R1 sequence. No relationship was found among genetic polymorphisms, identified by statistical analyses, and epidemiological or biological parameters. SIGNIFICANCE AND IMPACT OF THE STUDY: The analyses of five different genomic sequences of the 35 strains belonging to subgroup 16SrI-B allowed a finer distinction among them, confirming that the polymorphism level of 16S rDNA is too low to be adopted as unique parameter for classification.     

Cryptic species and systematics of the hynobiid salamanders of the Liua–Pseudohynobius complex: Molecular and phylogenetic perspectives

Using mitochondrial DNA sequencing and allozyme electrophoresis, we examined 18 populations of the Liua–Pseudohynobius complex, endemic to China. Based on their phylogenetic affiliation and exhibited fixed allelic differences, the complex comprises at least six species, two of which are previously unknown cryptic species. The complex is clearly divided into two groups, genus Liua including Liua shihi and Liua tsinpaensis, and genus Pseudohynobius including Pseudohynobius flavomaculatus, Pseudohynobius shuichengensis and the two new species. The previously often used genus name Ranodon is inappropriate, because the type species of the genus, Ranodon sibricus, is distantly related to this complex. The species diversity among Chinese hynobiid salamanders are far from being recognized and further effort should be directed at extensive field collection in central and western China.     

Application of rDNA-PCR amplification and DGGE fingerprinting for detection of microbial diversity in a Malaysian crude oil

Two culture-independent methods, namely ribosomal DNA libraries and denaturing gradient gel electrophoresis (DGGE), were adopted to examine the microbial community of a Malaysian light crude oil. In this study, both 16S and 18S rDNAs were PCR-amplified from bulk DNA of crude oil samples, cloned, and sequenced. Analyses of restriction fragment length polymorphism (RFLP) and phylogenetics clustered the 16S and 18S rDNA sequences into seven and six groups, respectively. The ribosomal DNA sequences obtained showed sequence similarity between 90 to 100% to those available in the GenBank database. The closest relatives documented for the 16S rDNAs include member species of Thermoincola and Rhodopseudomonas, whereas the closest fungal relatives include Acremonium, Ceriporiopsis, Xeromyces, Lecythophora, and Candida. Others were affiliated to uncultured bacteria and uncultured ascomycete. The 16S rDNA library demonstrated predomination by a single uncultured bacterial type by >80% relative abundance. The predomination was confirmed by DGGE analysis.     

中国小鲵科属、种分类变更的回顾

小鲵科为亚洲特有的有尾两栖动物,是现生有尾目10科中第三大科.中国是小鲵科动物属、种分布的主要区域,且研究其分类历史较久,学者较多,故导致学术观点差异较大.本文对中国小鲵科分类地位发生变更的部分属(巴鲵属Liua、拟小鲵属Pseudohynobius、肥鲵属Pachyhynobius)、种(秦巴巴鲵L. tsinpaensis、满洲小鲵Hynobius mantchuricus、豫南小鲵H. yunanicus、弱唇褶山溪鲵Batrachuperus cochranae和太白山溪鲵B. taibaiensis)的变更历史进行回顾,旨在为小鲵科动物的分类提供一个清晰的背景知识.     

Nucleic Acid Homologies Among Species of Saccharomyces

Evolutionary divergence among species of the yeast genus Saccharomyces was estimated from measurements of deoxyribonucleic acid (DNA)/DNA and ribosomal ribonucleic acid (RNA)/DNA homology. Much diversity was found in the DNA base sequences with several species showing little or no homology to the three reference species, S. cerevisiae, S. lactis, and S. fragilis. These three reference species also showed little or no homology to each other. On the other hand the diversity among ribosomal RNA base sequences was small since most species showed a high degree of homology to the reference species. The arrangement of species based on ribosomal RNA homologies agrees in most cases with current taxonomic groupings. A yeast hybrid (S. fragilis x S. lactis) was shown to contain two nonhomologous genomes. A minimum genome size of 9.2 x 10(9) daltons for S. cerevisiae was calculated from the rate of DNA renaturation.