贵阳常见嗜尸性蝇类生活史和蛹期转录组学研究

发布时间：2018-02-15 08:54

本文关键词： 法医昆虫学发育历期转录组基因差异表达　出处：《贵州医科大学》2017年硕士论文　论文类型：学位论文

【摘要】：目的:观察贵阳地区夏季瘦叶带绿蝇(Hemipyrellia ligurriens)、厩腐蝇(Muscina stabulans)、台湾别麻蝇(Boettcherisca formosensis)在室内自然条件下生活史与积温的关系;探讨棕尾别麻蝇(Boettcherisca peregrine)蛹3个不同发育时间转录组的差异表达,为进一步探究棕尾别麻蝇蛹各个时间转录组中的差异基因提供实验数据。方法:1.于贵州医科大学和贵州警官职业学院两地,分别放置家兔尸体诱捕台湾别麻蝇、厩腐蝇、瘦叶带绿蝇3种嗜尸性蝇类进行饲养直至产卵,观察3种嗜尸性蝇类在贵阳地区夏季自然条件下各发育阶段的历期并记录温、湿度。2.于贵州医科大学室外放置实验用家兔尸体,采集棕尾别麻蝇进行人工饲养。待其产下幼虫至化蛹后,每24 h收集一次蝇蛹,每次2粒,直至羽化出成虫,选取第1、5、10天收集的蝇蛹,提取总RNA测序进行文库构建,使用软件edgeR对基因差异表达进行分析,筛选差异基因做实时荧光定量验证,对已知基因(Unigene)进行京都数据库(KEGG)代谢通路分析,使用MISA对Unigene进行简单重复序列(SSR)检测。结果:1.台湾别麻蝇6月、7月、8月发育历期为17.00d、15.19d、15.7d;厩腐蝇6月、7月发育历期为16.13d、13.9d;瘦叶带绿蝇7月发育历为13.06d。其中厩腐蝇与瘦叶带绿蝇在7月发育历期相似;厩腐蝇6月、7月发育历期时间和台湾别麻蝇6月与7月、8月发育历期有差异。统计出3种嗜尸性蝇类的发育总历期和积温的基础数据。2.获得各分组样品基因组表达水平,共捕获38727条Unigenes,将不同时间的蝇蛹转录组结果构建数据库进行两两比较,差异基因总数为5283,将差异表达基因进行可视化散点图和火山图构建以及模式聚类构建,对差异基因GO分类统计和富集分析并将差异性明显的酪氨酸羟化酶(TH)基因与CBM-14基因筛选出来做实时荧光定量与测序结果一致。棕尾别麻蝇蛹转录组共有8856个Unigene被注释,被注释的代谢通路有336个。SSR查找发现,从Unigene中共查找到SSR位点占总比例的22.32%。其中,单核苷酸重复所占比例最高,其次是三核苷酸重复。SSR不同重复基元类型中,出现频率最高的为A/T,其次是AAC/GTT,AC/GT。结论:1.厩腐蝇、台湾别麻蝇、瘦叶带绿蝇的发育历期与温度变化有关,并获得三种嗜尸性蝇类的生活史数据。2.构建贵阳地区棕尾别麻蝇蛹3个不同发育时间的转录组数据库,用于寻找差异基因。3.TH基因与CBM-14基因在棕尾别麻蝇蛹的三个不同发育时间表达有显著差异。
[Abstract]:Objective: to observe the relationship between the life history and accumulated temperature of Hemipyrellia ligurriensus, Muscina stabulanssis and Boettcherisca formosensis in summer in Guiyang area, and to investigate the differential expression of three developmental time transcriptional groups of Boettcherisca peregrine pupae. In order to provide experimental data for further study on the differential genes in the pupae transcriptional groups at different times, methods: 1. The carcasses of rabbits were placed in Guizhou Medical University and Guizhou Police officers Vocational College respectively to trap and trap stables and stables of Ephedra tinctata and saproot fly, respectively, in Guizhou Medical University and Guizhou Academy of Police officers. Three species of necrophagous flies of the thin leaf belt were raised until they laid eggs. The development periods of the three species were observed and the temperature and humidity were recorded. 2. The experimental rabbits were placed in Guizhou Medical University outdoors. The pupae was collected once every 24 hours, until adult emergence. The pupae was collected from the pupae collected on day 1, 5 and 10, and the total RNA was sequenced to construct the library. The differential gene expression was analyzed by software edgeR, the differentially expressed genes were screened for real-time fluorescence quantitative verification, and the known gene edgeR was used to analyze the metabolic pathway in Kyoto database. MISA was used to detect the simple repeat sequence of Unigene. Results: 1. On June, July, August, the developmental period of Unigene was 17.00 dU 15.19 d (15.19 d), that of stable saproot fly was 16.13 d (13.9d) and that of thin leaf-belt green fly was 13.06 d. The developmental period of stables saproot and thin leaf was 13.06 d. The developmental period of green fly was similar in July. There were differences between the developmental periods of the stables saproot on June and July and the developmental periods of Elaeagnus chinensis on June and June, respectively. The basic data of the total developmental duration and accumulated temperature of the three species of necrophagous flies were analyzed. 2. The genomic expression levels of each group of samples were obtained. A total of 38727 Unigenes were captured, and the results of pupae transcriptome construction at different times were compared. The total number of differentially expressed genes was 5283. The differentially expressed genes were constructed by visual scatter map, volcanic map and pattern clustering. The differential gene go classification statistics and enrichment analysis, and the screening of different tyrosine hydroxylase THH gene and CBM-14 gene for real-time fluorescence quantitative analysis were consistent with the sequencing results. A total of 8856 Unigene were annotated in the pupae transcriptome of Magnus melanogaster. Of the 336 noted metabolic pathways, SSR loci were found to account for 22.32% of the total number of SSR loci from Unigene, followed by trinucleotide repeats, followed by trinucleotide repeats. The highest frequency of occurrence was A / T, followed by AAC / GTT / AC / GT.Conclusion: 1. The developmental period of stables saproot fly, Taiwan flax fly, thin leaf belt green fly is related to temperature change. The life history data of three species of necrophagous flies were obtained. 2. The transcriptome database of three different developmental periods of the pupae of Ephedra melanogaster was constructed in Guiyang area. There were significant differences in the expression of th gene and CBM-14 gene in the pupae of Magnus melanogaster at three different developmental times.
【学位授予单位】：贵州医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：D919

【相似文献】