中华蜜蜂细胞吞噬与包囊作用相关基因全长转录本鉴定及分析  

Identification and analysis of full-length transcripts of genes relative to phagocytosis and capsulation in Apis cerana cerana

作　　者：郭思佳 张凯遥 荆欣 高旭泽 冯佩林 邹培缘 张浩宇 陈大福 郭睿 付中民 GUO Sijia;ZHANG Kaiyao;JING Xin;GAO Xuze;FENG Peilin;ZOU Peiyuan;ZHANG Haoyu;CHEN Dafu;GUO Rui;FU Zhongmin(College of Animal Sciences(College of Bee Science),Fujian Agriculture and Forestry University,Fuzhou,Fujian 350002,China;Apitherapy Research Institute of Fujian Province,Fuzhou,Fujian 350002,China)

机构地区：[1]福建农林大学动物科学学院(蜂学学院),福建福州350002 [2]福建省蜂疗研究所,福建福州350002

出　　处：《西北农林科技大学学报（自然科学版）》2024年第3期1-10,共10页Journal of Northwest A&F University(Natural Science Edition)

基　　金：国家自然科学基金面上项目(32172792);国家现代农业产业技术体系建设专项资金项目(CARS-44-KXJ7);福建农林大学硕士生导师团队项目;福建农林大学动物科学学院(蜂学学院)科研扶持项目;福建省省级大学生创新创业训练计划项目(202310389027,X202310389084)。

摘　　要：【目的】系统鉴定和分析中华蜜蜂(Apis cerana cerana)吞噬与包囊作用相关基因和全长转录本,为深入开展相关基因和剪接体的功能研究奠定基础。【方法】基于前期已获得的高质量中华蜜蜂纳米孔长读段测序数据,通过Blast工具将全长转录本比对Nr数据库筛选出吞噬与包囊作用相关基因和全长转录本。利用gffcompare软件将全长转录本与东方蜜蜂(Apis cerana)参考基因组上注释的转录本进行比较,鉴定未注释的新基因和新转录本。利用TAPIS pipeline预测和分析吞噬与包囊作用相关基因的可变多聚腺苷酸化(alternative polyadenylation, APA)位点,并通过TBtools软件鉴定APA位点上游的基序(motif)。使用Astalavista软件鉴定可变剪接(alternative splicing, AS)事件,并通过IGV浏览器进行结构可视化。通过RT-PCR验证AS事件的真实性。【结果】共鉴定到中华蜜蜂吞噬与包囊作用相关的基因66个和全长转录本395条,发掘出东方蜜蜂参考基因组未注释的2个新基因和303条新转录本。对参考基因组已注释的34个基因进行了结构优化,分别延伸了18个基因的5′端和12个基因的3′端,同时延长了4个基因的5′端和3′端。共鉴定到含有1个及以上APA位点的吞噬与包囊作用相关基因47个,其中多于5个APA位点的基因最多,为32个。在APA位点上游鉴定到多个基序,一致性序列为:GRBGCNKSDAACAAYTRBGCBMRNGGBYAYTAYWCNVWNGG。共鉴定到吞噬与包囊作用相关基因的AS事件296次,其中包括131次可变3′端剪接(alternative 3′splice site, A3SS)、85次内含子保留(intron retention, IR)、70次可变5′端剪接(alternative 5′splice site, A5SS)和10次外显子跳跃(exon skipping, ES)。RT-PCR结果显示,扩增的目的片段大小符合预期,证实了随机选择的2次AS事件的真实性。【结论】系统鉴定了中华蜜蜂吞噬与包囊作用相关基因和全长转录本以及AS事件和APA位点,优化了东方蜜蜂参考�【Objective】Systematic identification and investigation of genes and full-length transcripts associated with phagocytosis and capsulation in Apis cerana cerana were conducted,aiming to provide basis for further functional study on related genes and isoforms.【Method】Based on previously gained high-quality Nanopore long read sequencing data from Apis cerana cerana,the mapping of full-length transcripts to Nr database was conducted by Blast tool to identify full-length transcripts relative to phagocytosis and capsulation.The full-length transcripts were compared with those annotated on reference genome using the gffcompare software to identify unannotated novel genes and full-length transcripts.Prediction and analysis of alternative polyadenylation(APA)sites were conducted with TAPIS pipeline,followed by identification of motifs upstream of APA sites by TBtools software.Astalavista software was employed to identify alternative splicing(AS)events followed by structural visualization by IGV browser.RT-PCR was performed to validate the authenticity of AS events.【Result】A total of 66 genes and 395 full-length transcripts relevant to phagocytosis and capsulation in Apis cerana cerana were discovered,including 2 new genes and 303 new transcripts unannotated on the reference genome of Apis cerana.The structure of 34 annotated genes of the reference genome was optimized,among which 5′ends of 18 genes and 3′ends of 12 genes were extended,and 5′ends and 3′ends of 4 genes were prolonged.Additionally,47 genes related to phagocytosis and capsulation were identified to contain one or more APA sites.The number of genes with more than 5 APA sites was 32.Multiple motifs were identified in upstream of APA sites,and the consistent sequence was GRBGCNKSDAACAAYTRBGCBMRNGGBYAYTAYWCNVWNGG.In total,296 AS events were identified,including 131 alternatives 3′splice sites(A3SS),85 intron retention(IR),70 alternative 5′splice sites(A5SS)and 10 exon skipping(ES).RT-PCR showed that the sizes of amplified fragments were con

关 键 词：中华蜜蜂 吞噬作用 包囊作用 全长转录本 纳米孔测序 可变剪接 可变多聚腺苷酸化 

分 类 号：S895.137[农业科学—特种经济动物饲养]