我国32种鸟类DNA条形码分析

DNA条形码是一种分子分类方法,近年来在物种鉴定方面得到迅速的发展和应用.本研究分析了我国27属32种鸟类(61只)的线粒体细胞色素c氧化酶亚基Ⅰ(COⅠ)基因的条形码片段,分别用阈值法、聚类法和诊断核苷酸进行了分析,探究DNA条形码鉴定我国鸟类的准确性.结果显示,种内CO Ⅰ序列变异很小,种间存在较多的变异位点,种间的遗传距离显著大于种内的遗传距离,DNA条形码序列能够鉴定所有鸟类.     

基于DNA条形码的四川老君山国家级自然保护区尺蛾科昆虫种类鉴定

为了解四川老君山国家级自然保护区鳞翅目Lepidoptera尺蛾科Geometridae资源,于2019年6—8月和2020年5—6月通过灯诱法采集2402号标本,运用DNA条形码技术对标本进行分子鉴定.通过BOLD数据库,邻接法构建系统发育树和ABGD在线工具对获得的297条线粒体细胞色素C氧化酶亚基Ⅰ(COⅠ)基因...     

斑腿蝗科Catantopidae七种蝗虫线粒体COⅠ基因的DNA条形码研究

以我国常见的斑腿蝗科Catantopidae7种蝗虫为对象测定了COⅠ基因序列,探讨COⅠ基因作为DNA条形码在识别蝗虫物种方面的可行性。结果表明,斑腿蝗科3属7种的DNA分类和形态学分类基本一致,该基因可以探讨蝗虫属、种分类单元的系统发育问题,为将线粒体基因组的COⅠ基因作为蝗虫DNA条形码进行分类鉴定手段的可行性提供一定的参考。     

三化螟COⅠ序列分析及DNA条形码研究

三化螟Scirpophaga incertulas是水稻的重要害虫之一。本研究通过对三化螟3个地理种群线粒体COⅠ基因的序列分析,初步探讨了中国三化螟种群间的遗传分化。36个三化螟样本共得到了9个单倍型,其中有17个变异位点,占总位点数的3.13%。系统进化分析表明,来源于各地理种群的单倍型散布在不同的进化支上,缺乏明显的地理分布格局。联合网上公布的相关物种序列进行的遗传分析表明,线粒体COⅠ基因能够对三化螟进行准确的物种鉴定。     

DNA条形码技术在田间常见蓟马种类识别中的应用

蓟马类害虫种类多、 体型小, 传统的形态学鉴定方法难以快速准确识别。本研究利用DNA条形码通用型引物, 以我国田间常见的25种蓟马为靶标扩增其线粒体DNA细胞色素C氧化酶亚基Ⅰ (mitochondrial cytochrome c oxidase subunit Ⅰ gene, mtDNA COⅠ) 基因 (约650 bp), 通过对靶标片段碱基序列的测序及比对分析, 以邻接法 (NJ法) 构建系统发育树, 并以Kimura双参数模型计算种内、 种间遗传距离。结果表明： 聚类分析与形态学鉴定结果一致, 表现为较长的种间分支和较短的种内分支, 每个单系分支对应一个物种, 同一物种不同单倍型的最初分支自展值均为100%。25种蓟马的种内平均遗传距离为0.0027, 种间平均遗传距离为0.2757, 种间遗传距离为种内遗传距离的102.1倍; 而且种内、 种间遗传距离没有重叠区域。结果说明基于COⅠ基因的DNA条形码技术可以用于不同种类蓟马的快速准确鉴别。     

DNA条形码识别Ⅰ.DNA条形码研究进展及应用前景

DNA条形码(DNA Barcoding)是近年来生物分类学中引人注目的发展热点.本文综述了DNA条形码的发展历史、识别原理以及公共数据库,并讨论了DNA条形码在检疫检验领域的应用前景.DNA条形码与DNA芯片技术的结合,将推动传统物种鉴定方法的更新,可在检疫检验领域中实现非专家检定,这对进出口口岸生物监测具有重要的理论意义和应用价值.     

基于线粒体COⅠ的DNA 条形码技术在贻贝科种类鉴定中的应用

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基于进化树、距离和特征的DNA条形码方法研究——以百花山地区草螟科为例

DNA条形码是一种快捷高效的分子鉴定新技术,近年来在动物分类学领域中得到迅速的发展和应用。在条形码的研究中有基于进化树、距离和特征3种常用的分析方法:第1种方法需要构建系统发育树,分析样本在树上的聚类情况;第2种方法依赖于物种种内和种间的序列差异;第3种则是通过一系列的诊断特征位点来鉴定物种。本研究扩增了北京百花山地区14种草螟科昆虫88个样本的线粒体细胞色素C氧化酶亚基Ⅰ(mitochondrial cytochrome c oxidase subunitⅠ,COⅠ)基因片段,分别基于进化树、距离和特征方法进行了分析,以探讨不同DNA条形码方法在草螟科物种鉴定中的可行性。结果表明:在使用邻接法(neighbor-joining,NJ)构建的系统发育树上,14个草螟物种各自聚成一个单系,均被成功区分。基于Kimura双参数模型计算遗传距离得出,种内和种内有一个明显的"barcoding gap",且ABGD软件对样本的划分完全符合形态鉴定结果。在所有的草螟物种中都找到了诊断核苷酸位点,基于特征来鉴定草螟物种的成功率为100%。结果显示,这3种方法对于本研究中所涉及的草螟都具有较好的区分,基于COⅠ基因的DNA条形码可以作为一种有效的工具在草螟科昆虫的物种鉴定中进行应用。     

中国果实蝇属种类的DNA条形码鉴定(双翅目,实蝇科)

将实验获得的25种果实蝇的155条COⅠ条形码序列,利用MEGA4.1的Kimura-2-Parameter模型进行了遗传距离分析和构建系统发育树,来检验线粒体COⅠ基因条形码序列对果实蝇属种类鉴定的有效性。研究表明COⅠ条形码序列能够对除桔小实蝇复合体外的中国果实蝇属种类进行准确鉴定。     

基于线粒体COⅠ基因的齿小蠹属昆虫DNA条形码研究

齿小蠹属（鞘翅目： 小蠹科）昆虫是植物检疫中经常截获的类群, 为探讨线粒体细胞色素C氧化酶亚基Ⅰ(COⅠ)基因的特定区段作为DNA条形码快速准确鉴定齿小蠹种类的可行性, 以齿小蠹属昆虫为研究对象, 测定分析了线粒体COⅠ基因462 bp碱基序列。序列分析结果显示： 变异位点为259个, 保守位点203个, 简约信息位点181个, 自裔位点78个。所有位点中, A, G, C和T碱基平均含量分别为30.7%, 16.5%, 17.0%和35.8%。A+T含量较高, 为66.5%, 明显高于G+C含量, 表现明显的A+T碱基偏嗜, 且A与T含量相当, 符合昆虫线粒体基因碱基组成的基本特征。转换与颠换结果显示： 该段序列未达到饱和, 可以得到准确的进化分析。利用Kimura 2-parameter模型分析遗传距离得到, 同物种间的遗传距离介于0.002~0.007之间, 不同种间的遗传距离介于0.056~0.431间, 平均遗传距离为0.199, 说明该段序列能够区分不同物种。基于COⅠ基因序列构建的邻接法系统发育树（NJ树）显示, 同一物种聚为同一小支, 且分支自展值均为100%; 近缘种能聚集在一起, 且置信度很高（≥97%）。结果表明应用基于COⅠ基因片段的DNA条形码进行齿小蠹属昆虫分类鉴定具有可行性。     

RECOGNIZING DINOFLAGELLATE SPECIES USING ITS rDNA SEQUENCES1

Dinoflagellate taxonomy is based primarily on morphology and morphometric data that can be difficult to obtain. In contrast, molecular data can be rapidly and cost‐effectively acquired, which has led to a rapid accumulation of sequence data in GenBank. Currently there are no systematic criteria for utilizing taxonomically unassigned sequence data to identify putative species that could in turn serve as a basis for testable hypotheses concerning the taxonomy, diversity, distribution, and toxicity of these organisms. The goal of this research was to evaluate whether simple, uncorrected genetic distances (p) calculated using ITS1/5.8S/ITS2 (ITS region) rDNA sequences could be used to develop criteria for recognizing putative species before formal morphological evaluation and classification. The current analysis used sequences from 81 dinoflagellate species belonging to 14 genera. For this diverse assemblage of dinoflagellate species, the within‐species genetic distances between ITS region copies (p=0.000–0.021 substitutions per site) were consistently less than those observed between species (p=0.042–0.580). Our results indicate that a between‐species uncorrected genetic distance of p≥0.04 could be used to delineate most free‐living dinoflagellate species. Recently evolved species, however, may have ITS p values <0.04 and would require more extensive morphological and genetic analyses to resolve. For most species, the sequence of the dominant ITS region allele has the potential to serve as a unique species‐specific “DNA barcode” that could be used for the rapid identification of dinoflagellates in field and laboratory studies.     

中国蜜环菌生物种的RAPD分析

以中国蜜环菌(Armillaria mellea (vahl: Fr.) Kummer)生物种B．生物种E分别与欧洲种A. gallica 和A. ostoyae交配,用AP—PCR技术分析了中国五个生物种及欧洲2个种的代表菌株的系统发育关系。根据系统聚类分析的结果,将其分为4个群：生物种B与A．Gallia, A与C,D与E,A. ostoyae 单独为一群。这与交配试验及RAPD图谱直观分析的结果一致,最小支撑树也支持将中国生物种B鉴定为A．Gallica。证明RAPD是研究蜜环菌生物种的进化关系的有用手段。     

WILT OF LUCERNE CAUSED BY SPECIES OF VERTICILLIUM

The incidence and symptoms of a disease of lucerne induced by Verticillium spp. are much less severe immediately after the crop has been first cut than later when the crop has matured. Infection becomes more marked as the season advances and the age of the ley increases, and cuts taken in the third harvest year are proving unprofitable.
In cross-inoculation studies the causal organisms of lucerne wilt, V. albo-atrum and V. dahliae prove to be only mild pathogens to hop, potato, tomato and antirrhinum, and are not pathogenic to sainfoin and clover. The last two may therefore be cultivated as fodder crops in those areas most severely affected by lucerne wilt, but the cultivation of potatoes after lucerne should be discouraged, particularly if lucerne is to be sown in future years, since the potatoes might build up a reservoir of inoculum of the lucerne pathogen. In studies of host-parasite relations it is shown that V. albo-atrum and V. dahliae from lucerne, V. albo-atrum from hop, V. tricorpus from tomato and V. nigrescens and V. nubilum from potato vary in their effects upon lucerne seedlings growing on agar media, V. albo-atrum from lucerne being, as in the field, the most virulent and the only one to reach the xylem vessels under the conditions of the experiments.
The dressings Agrosan-GN (containing mercury) and Fernasan (containing thiram) are toxic both to spores of Verticillium on lucerne seed coats and to infected plant material carried with seed, and although they are also toxic in vitro to Rhizobium mellelotus this did not prevent dressed seeds which had been inoculated with the bacterium from developing into healthy seedlings bearing normal root-nodules.     

WILT OF LUCERNE CAUSED BY SPECIES OF VERTICILLIUM

A wilt disease of lucerne caused by species of Verticillium is described: at twenty-eight disease areas in England and Wales the pathogen was V. albo-atrum , while at one site it was V. dahliae . Both pathogens form superficial conidia on the basal regions of infected stems. It is shown that V. albo-atrum is introduced into a new area in contaminated plant material harvested with the seed from an infected crop. Rapid secondary spread of the disease follows the dissemination of spores from infected stems, and by contact of these and transported fragments of diseased tissues with the wounded surfaces of recently cut lucerne plants. It is recommended that seed should be collected only from healthy crops and that machinery, footwear, etc. should be disinfected before leaving a site of infection.
Manurial trials showed that the incidence of wilt induced by V. albo-atrum was very severe under all soil conditions tested, whereas V. dahliae is a virulent pathogen only to plants in soil rich in superphosphate. Generally the more vigorous the growth of the lucerne–in soils rich in potash and hoof and horn–the more rapid is both the onset of wilt and the resulting secondary spread of the disease throughout the crop.     

THE SPREAD OF DISEASES CAUSED BY SPECIES OF VERTICILLIUM

An investigation has been made of the spread of Verticillium disease of tomato and antirrhinum by root contact between diseased and healthy plants, and by the growth of the fungi through the soil. V. alboatrum and V. Dahliae spread rapidly by root contact, while V. tricorpus, V. nigrescens and V. nubilum showed no appreciable spread. Where the tomato and antirrhinum hosts wilted and died the causal organism was invariably isolated from the soil near the roots of the dead plants, but where the inoculated host harboured species without showing disease symptoms, as in the case of V. tricorpus and V. nubilum in antirrhinum, the parasite was never obtained from the soil. Moreover, these species in antirrhinum, and V. nigrescens and V. nubilum in tomato would appear to be mild pathogens since when they were already well established V. alboatrum was later actually found to enter and parasitize the respective hosts and bring about its own typical disease symptoms.
Spread through the soil in the absence of hosts was tested by the insertion into it of infected wheat grains followed by later attempts to isolate the fungi at varying distances from the point of infection. V. alboatrum, V. Dahliae and V. tricorpus showed little or no growth through the soil, whereas V. nigrescens and V. nubilum spread saprophytically to some extent, though never attaining sufficient concentration to cause appreciable infection of later planted tomatoes.
The conclusion is reached that spread of disease caused by species of Verticillium takes place mainly by root contact, with a rapidity relatively proportional to that of the death of the host. This finding, together with the lack of spread from antirrhinum plants infected with the non-lethal V. tricorpus and V. nubilum , suggests that the parasites remain in the vascular tracts until the death of the hosts.     

五种水螅基因组DNA的RAPD多态性研究

利用RAPD技术对水螅科2属5种水螅进行了基因组DNA多态性研究.80个随机引物筛选出18个引物,共扩增出303个清晰稳定的DNA片段,分子量在0.3～3.0kb之间.以UPGMA聚类法构建树状图,阐明水螅属种的分类关系:大庆水螅Hydra daqingensis、北京水螅H.beijingensis和强壮水螅H.robusta 3种分成两组并聚在一起;不同属的漠河水螅H.mohensis和绿水螅Chlorohydra sp.聚为一支;由此2属5种水螅分为两个支系.     

原位复染鉴别细胞的凋亡与坏死

用10μg／ml去甲斑蝥素作用于人红白血病K562细胞24h,诱导其发生凋亡。采用台盼蓝染液染色,再瑞特-吉姆萨染液原位复染的方法,验证了凋亡细胞与坏死细胞质膜通透性不同的说法。利用此方法可以在普通光学显微镜下把一张涂片上的凋亡细胞、坏死细胞及活细胞区别开来。该实验也表明,常规用台盼蓝染液拒染细胞数来统计活细胞数的方法是值得商榷的。