法医学杂志

法医学杂志 ›› 2021, Vol. 37 ›› Issue (3): 318-324，331.DOI: 10.12116/j.issn.1004-5619.2020.401214

所属专题： 法医昆虫学

• 专题 • 上一篇    下一篇

不同发育时期大头金蝇蛹转录组测序分析

王启燕， 张红玲， 任峥， 刘玉波， 季晶焱， 黄江   

  1. 贵州医科大学法医学院 法医司法鉴定中心，贵州 贵阳 550004
  • 发布日期:2021-06-25 出版日期:2021-06-28
  • 通讯作者: 黄江，女，教授，博士研究生导师，主要从事法医物证学、法医昆虫学教学、科研及鉴定；E-mail：mmm_hj@126.com
  • 作者简介:王启燕（1988—），女，硕士，讲师，主要从事法医昆虫学、法医物证学研究；E-mail：136713438@qq.com
  • 基金资助:
    贵州省高层次“百”层次创新型人才资助项目（黔科合平台人才[2020]6012）；贵州省高技术产业化示范工程资助项目[黔发改高技（2016）1345号]；贵州省科技计划资助项目（黔科合支撑[2019]2825号，黔科合支撑[2020]4Y057号）；贵州省科技计划资助项目（黔科合[2020]1Y353号）；贵州省科技厅计划资助项目（黔科合LH字[2016]7360）

Transcriptome Sequencing Analysis of Chrysomyia Megacephala Pupae in Different Growing Periods

WANG Qi-yan, ZHANG Hong-ling, REN Zheng, LIU Yu-bo, JI Jing-yan, HUANG Jiang#br#   

  1. Department of Forensic Medicine, School of Forensic Medicine, Guizhou Medical University, Guiyang 550004, China
  • Online:2021-06-25 Published:2021-06-28

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摘要： 目的 研究大头金蝇蛹的发育调控、环境适应、表观调控等问题，以获得不同发育时期转录组数据，为法医学应用奠定基础。 方法 饲养大头金蝇，待其化蛹，从开始化蛹至羽化出成虫，每24 h收集1次，每次收集3粒蝇蛹，-80 ℃保存备用。采用Illumina Hiseq4000进行高通量测序，获得的非重复序列基因（Unigene），使用NCBI比对工具BLAST分别与NR、STRING、SWISS-PROT瑞士蛋白质数据库（包括Pfam）、GO数据库、COG数据库、KEGG数据库进行比对，获得相应的注释信息，采用每百万测序碱基中基因外显子每千个碱基长度中所包含的测序片断数（fragments per kilobase of exon model per million mapped reads，FPKM）方法计算测序所得Unigene在6个不同发育时期大头金蝇蛹的表达量，并以不同发育时期FPKM表达量比值的log2倍数绝对值大于1（即log2|FC|>1）及错误发现率小于0.05为标准进行差异基因筛选。 结果 平均温度为25.6 ℃时，大头金蝇蛹从开始化蛹至羽化出成虫历时6 d，共获得43 408条Unigene，平均长度为905 bp，在NR、SWISS-PORT、Pfam、STRING、KEGG数据库分别有32 500、18 720、13 542、9 191、18 720条Unigene获得注释，将不同发育时期蛹两两比较进行差异基因分析，差异表达基因数有801~5 307个，差异表达基因总数45 676个。 结论 大头金蝇蛹转录组数据在不同发育时期基因表达具有差异性，为进一步探究嗜尸性蝇蛹各个时期转录组变化提供了良好的基础，为挖掘大头金蝇蛹发育相关基因提供依据。

关键词: 法医昆虫学, 法医遗传学, 死亡时间, 测序, 转录组, 大头金蝇

Abstract: Objective To study the growth regulation, environmental adaption and epigenetic regulation of Chrysomyia Megacephala pupae, in order to obtain the transcriptome data of Chrysomyia Megacephala in different growing periods, and lay the foundation for forensic application. Methods The Chrysomyia Megacephala was cultivated and after pupation, 3 pupae were collected every 24 h from pupation to emergence, and stored at -80 ℃ for later use. High-throughput sequencing was performed by Illumina Hiseq 4000 and Unigenes were obtained. The Unigenes were compared by comparison tool BLAST from NCBI in databases such as NR, STRING, SWISS-PROT （including Pfam）, GO, COG, KEGG in order to obtain the corresponding annotation information. The expression amount of Unigenes obtained by sequencing in Chrysomyia Megacephala in six different growing periods was calculated by FPKM method, and the discrepant genes were screened according to the following standards： the log2 multiple absolute value of FPKM expression amount between two different growing periods must be larger than 1 （log2|FC|>1）, and the false discovery rate must be less than 0.05. Results When the mean temperature was 25.6 ℃, Chrysomyia Megacephala emerged 6 d after they pupated. A total of 43 408 pieces of Unigenes were obtained and their mean length was 905 bp, of which 32 500, 18 720, 13 542, 9 191 and 18 720 pieces were annotated by NR, SWISS-PORT, Pfam, STRING and KEGG databases. According to the discrepant gene analysis of pupae in two different growing periods, the number of genes with variants ranged from 801 to 5 307, and the total number of discrepant genes was 45 676. Conclusion The gene expressions of the transcriptome data of Chrysomyia Megacephala pupae in different growing periods are different. The results provided a good foundation for further research on the transcriptome changes in each period of the pupae of sarcosaprophagous flies and provided the basis for exploring the genes associated with the growth of Chrysomyia Megacephala pupae.

Key words: forensic entomology, forensic genetics, postmortem interval, sequencing, transcriptome, Chrysomyia Megacephala

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