图兰扇头蜱线粒体基因组序列分析及其种内鉴定新基因标志物的建立

发布时间：2018-05-15 18:22

本文选题：图兰扇头蜱 + 线粒体基因组　；参考：《石河子大学》2017年硕士论文

【摘要】：目的:(1)对图兰扇头蜱线粒体全基因组序列进行测定并对其结构特点进行分析,探讨扇头蜱属系统发育关系。(2)设计3个新的图兰扇头蜱种内鉴定的基因标志物,对图兰扇头蜱进行种内分子鉴定。方法:(1)根据Gen Bank数据库的扇头蜱属线粒体全基因组序列,设计出6对线粒体全基因组引物用于PCR扩增中国新疆图兰扇头蜱线粒体全基因组序列。运用多个基因分析软件进行线粒体全基因组序列的组装、分析,并进行进一步的12S r DNA和cox1基因的最大似然法(Maximum Likelihood,ML)系统发生学分析。(2)采集中国新疆南北疆7个县市等地区212只图兰扇头蜱,阿尔巴尼亚4个县市103只图兰扇头蜱,结合蜱的形态学特征及蜱线粒体16S r RNA和cox1基因,对所采集的蜱进行蜱种的初步形态学及分子生物学鉴定并进行ML法系统发育分析。根据扇头蜱属线粒体全基因组中的保守区域,设计3个新的线粒体序列基因标志物—nad1-16S r RNA(简称为N1),nad2-cox1(简称为N2),cox1-t RNA-Lys(简称为C1),扩增和分析中国新疆和阿尔巴尼亚图兰扇头蜱的N1、N2、C1片段并进行种内基因差异分析和ML法系统发生学分析。结果:(1)在国际上首次测出图兰扇头蜱全线粒体基因组长为14717 bp,其由13个蛋白质编码基因、22个t RNA基因、2个核糖体RNA基因和2个非编码区组成,各基因排列顺序与后沟类硬蜱相似。图兰扇头蜱全线粒体基因组核苷酸组成为A=37.80%,T=40.00%,G=9.90%,C=12.30%,A+T含量为77.80%;ML系统进化树显示图兰扇头蜱位于后沟类硬蜱分枝上,且与中国血红扇头蜱亲缘关系较近,其和血红扇头蜱中国株一道,与血红扇头蜱美国株形成姊妹种关系。(2)本研究的3个新的基因标志物(N1、N2、C1)是有效的图兰扇头蜱种内鉴定的基因工具,这3个基因标志物在中阿两国的图兰扇头蜱的差异分别为3.57-4.07%、3.57-4.39%和3.18-4.69%,将中国和阿尔巴尼亚的图兰扇头蜱分为两个不同的亚种。结论:(1)本研究首次在国际上完成图兰扇头蜱线粒体的全基因组测序;与大多数典型的后生动物线粒体基因组类似,图兰扇头蜱线粒体基因组包含37个基因,包括13个蛋白质编码基因、2个r RNA基因和22个t RNA基因且仅有一个拷贝,具备2个非编码控制区,呈闭合环状结构。就目前的研究和数据而言,本研究的图兰扇头蜱与血红扇头蜱中国株在系统发生关系上最近,但未来仍然需要扩增扇头蜱属的其他种的mt DNA来进行更广阔的分析。(2)本研究设计的3个新的图兰扇头蜱线粒体基因标志物—N1、N2、C1是有效的图兰扇头蜱种内鉴定基因标志物,将中国新疆和阿尔巴尼亚的图兰扇头蜱成功分为2个不同的亚种进化枝。图兰扇头蜱至少存在两个不同的亚种,未来应该大量采集世界其他地域的图兰扇头蜱进行亚种的研究。
[Abstract]:Objective to determine the whole mitochondrial genome sequence of Tulan tick and analyze its structural characteristics, and to study the phylogenetic relationship. The intraspecific molecular identification of ticks was carried out. Methods according to the whole mitochondrial genome sequence of Gen Bank database, six pairs of mtDNA primers were designed to amplify the whole mitochondrial genome sequence of Tulan tick in Xinjiang, China. The whole mitochondrial genome sequence was assembled and analyzed by multiple gene analysis software. Further phylogenetic analysis of 12s r DNA and cox1 genes by Maximum Likelihoodtus (MLL) phylogenetic analysis was carried out. 212 Tulan head ticks were collected from 7 counties and cities in Xinjiang, China, and 103 from 4 counties in Albania. Combined with the morphological characteristics of ticks and the 16s r RNA and cox1 genes of ticks mitochondria, the preliminary morphological and molecular biological identification and phylogenetic analysis of the collected ticks were carried out by ML method. According to the conserved region of the mitochondrial genome of the genus Acarus, To design three new mitochondrial gene markers -nad1-16S r RNA-nad2-cox1 (abbreviated as N2C2Cox1-t RNA-Lys-Lys-Lys1), amplify and analyze the N _ 1N _ 2N _ 2N _ 2C _ 1 fragment of ticks from Xinjiang, China and Albania, and carry out intraspecific gene differential analysis and ML. Analysis of phylogeny of legal system. Results the length of the whole mitochondrial genome was 14717 BP for the first time in the world. It was composed of 13 protein coding genes, 22 t RNA genes, 2 ribosomal RNA genes and 2 non-coding regions. The sequence of genes was similar to that of Hypodes. The nucleotide composition of the whole mitochondrial genome of Trichophynchus thuringii is A37.800.0.0.The content of Cn12.30AT is 77.800.The phylogenetic tree shows that the tick is located on the branches of the posterior sulcus and is closely related to the Chinese Haematopsis, which is associated with the Chinese strain of Haemophorus. In this study, three new genetic markers, N _ (1) N _ (2) C _ (1), are effective genetic tools for intraspecific identification. The difference of the three gene markers between China and Albania was 3.57-4.073.57-4.39% and 3.18-4.69respectively. The Chinese and Albanian ticks were divided into two different subspecies. Conclusion the whole mitochondrial genome of Tulan head tick was sequenced in the world for the first time in this study, which was similar to that of most typical metazoa. The mitochondrial genome of Tulan head tick contained 37 genes. There are 13 protein coding genes, 2 r RNA genes and 22 t RNA genes with only one copy. There are 2 non-coding control regions with closed ring structure. As far as the current research and data are concerned, the phylogenetic relationship between the Chinese strains of Tulan tick and Hematoptera pyramidis is the most recent in this study. But in the future, mt DNA of other species of the genus Thalanopsis need to be amplified for a broader analysis.) the three new mitochondrial gene markers -N1N _ 2N _ 2C _ 1 designed in this study are effective markers for intraspecific identification of ticks. Two subspecies evolutionary branches were successfully divided into two subspecies from Xinjiang, China and Albania. There are at least two different subspecies in Tulan tick. In the future, a large number of species should be collected from other parts of the world.
【学位授予单位】：石河子大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R384.4

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