苍蝇群落演替及MtDNA分子标记的法医学研究

发布时间：2018-07-28 16:32

【摘要】：目的研究呼和浩特地区夏秋季常见嗜尸性苍蝇生态群落组成与演替;利用线粒体DNA(mtDNA)上细胞色素氧化酶辅酶Ⅰ和Ⅱ(COⅠ和COⅡ)以及16SrDNA中278bp和635bp以及513bp基因序列,鉴定嗜尸性苍蝇的种类,解决依据形态学方法不能鉴定苍蝇卵的种类、很难鉴定幼虫、蛹种类的难题,为法医鉴别嗜尸性苍蝇及其幼虫、蛹、卵种类提供依据。同时,探讨嗜尸性苍蝇mtDNA基因序列分析所得的遗传距离与所处地理纬度变化的相关与回归关系,为法医判断嗜尸性苍蝇所属地理种群,进一步准确鉴定其种类、推断死亡时间和地点创造条件。方法连续两年用观察法研究7～10月份,家兔、狗尸体、鸡、鱼内脏等试验材料上的嗜尸性苍蝇生态群落组成与演替;随机采集放置在呼和浩特及成都地区室外草地家兔尸体上的嗜尸性苍蝇、幼虫、蛹、苍蝇腹中的卵。利用改进的小型昆虫DNA匀浆提取方法提取上述样本mtDNA通过Perkin-Elmer 9600扩增仪进行PCR扩增;琼脂糖水平电泳以及聚丙烯酰胺非变性凝胶连续缓冲体系垂直电泳和银染显色技术进行扩增结果检测;PCR胶回收试剂盒纯化;ABI 377测序仪测序;DNAMAN 4.0序列分析软件,进行序列比对,截取等长度片段;MEGA2.1软件包进行序列分析和构建系统发育树。同时,通过GenBank中BLAST检索系统检索世界性研究最为广泛的23个果蝇mtDNA上COⅠ中278bp基因序列,以及包括本试验组检测并上传的采集于呼和浩特和成都地区及世界其它地区的10个丝光绿蝇和10个铜绿蝇的mtDNA上COⅡ中的635bp基因序列;用MEGA2.1软件包进行序列分析和构建系统发育树,并将分析所得的各苍蝇间的遗传距离与地理纬度差运用SPSS统计软件进行回归和相关分析。结果该地区上述尸体上出现的双翅目嗜尸性苍蝇3科8属10种,主要
[Abstract]:Objective to study the ecological community composition and succession of the common cadaver flies in summer and autumn in Hohhot area, and to identify the species of corpse flies by using mitochondrial DNA (mtDNA) cytochrome oxidase coenzyme I and II (CO I and CO II) and 278bp and 635bp and 513bp gene sequences in 16SrDNA to solve the failure of identification of flies according to the morphological method. The species of eggs is difficult to identify the difficult problem of the larva and the species of pupae. It provides a basis for the forensic identification of the cadaver flies and their larvae, pupae and eggs. At the same time, the correlation and regression relationship between the genetic distance and the geographical latitude of the mtDNA gene of the cadaver fly is discussed. Step by step accurately identify its species and deduce the conditions of death time and place creation.
Methods the ecological community composition and succession of the autopsy flies on the experimental materials such as 7~10 months, rabbits, dog bodies, chicken and fish viscera were observed for two years for 7~10 consecutive years, and the autopsy flies, larvae, pupae and flies in the abdomen of the rabbit bodies in Hohhot and Chengdu area were collected randomly. The improved small insect was used to make use of the improved small insect DNA. MtDNA was amplified by Perkin-Elmer 9600 amplification by homogenate extraction. The agarose level electrophoresis and polyacrylamide gel continuous buffer system vertical electrophoresis and silver staining technique were used to detect the amplification result, PCR gel recovery kit was purified, ABI 377 sequencer was sequenced, and DNAMAN 4 sequence analysis was soft MEGA2.1 software package carries out sequence analysis and constructs a phylogenetic tree. At the same time, the BLAST retrieval system in GenBank is used to retrieve the 278bp gene sequence in CO I of 23 fruit flies, the most widely studied fruit fly mtDNA, as well as the testing and uploading of this test group in Hohhot and Chengdu. The 635bp gene sequences in the mtDNA CO II of 10 mercerizing flies and 10 aeruginosa flies in the region and other regions of the world; the sequence analysis and construction of the phylogenetic tree were carried out by the MEGA2.1 software package, and the genetic distance between the flies was analyzed by the regression and correlation analysis using the SPSS statistical soft parts of the geographical latitude difference.
Results 3 species, 8 genera and 10 species of Diptera fly were found in the above bodies.
【学位授予单位】：四川大学
【学位级别】：博士
【学位授予年份】：2005
【分类号】：D919

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