基于转录组研究MPEF对毕赤酵母的致死机理  

Transcriptomic Investigation of the Mechanism of the Lethal Effect of Microchip Pulse Electric Field on Pichia

作　　者：朱宁[1] 于宁 朱月[1] 韦玉龙 张嘉颖 孙爱东[1] ZHU Ning;YU Ning;ZHU Yue;WEI Yulong;ZHANG Jiaying;SUN Aidong(College of Biological Sciences and Biotechnology,Beijing Forestry University,Beijing 100083,China)

机构地区：[1]北京林业大学生物科学与技术学院,北京100083

出　　处：《食品科学》2019年第4期130-137,共8页Food Science

基　　金：国家自然科学基金面上项目(31471593);"十三五"国家重点研发计划重点专项(2016YFD0400302);林业公益性行业科研专项(201504606)

摘　　要：探究微芯片脉冲电场(microchip pulse electric field,MPEF)技术对毕赤酵母的致死效果和致死机理。在400 V电压和80个脉冲条件下,MPEF技术可以很好地钝化毕赤酵母。借助新一代高通量测序技术对MPEF处理前后毕赤酵母基因的表达水平进行分析,将差异表达基因在基因本体(gene ontology,GO)和京都基因与基因组百科全书(Kyoto Encyclopedia of Genes and Genomes,KEGG)数据库中进行比对。测序结果表明,MPEF组和对照组差异变化显著的基因共794个,其中上调基因361个,下调基因433个;在MPEF作用下,半胱天冬蛋白酶活性、丙酮酸转运等显著上调,而核苷酸代谢、核糖体生物合成、蛋白质转录翻译、RNA聚合酶活性、超氧化物歧化酶活性、跨膜运输调节、维生素代谢等显著下调。采用实时荧光定量聚合酶链式反应对6个差异基因进行验证,得到与转录组测序结果一致的基因表达量变化趋势。毕赤酵母体内细胞结构的破坏、基因损伤、蛋白质合成和功能受限以及酶活性的改变可能是MPEF处理导致其死亡的主要原因。In this work,the effect of microchip pulse electric field(MPEF)treatment on inactivating Pichia and the underlying mechanism were investigated.A good inactivation of Pichia was achieved after 80 pulses at 400 V.The differentially expressed genes of Pichia before and after MPEF treatment were analyzed using new generation of highthroughput sequencing,and they were aligned to the gene ontology(GO)and the Kyoto Encyclopedia of Genes and Genomes(KEGG)databases.Results showed that a total of significantly differentially expressed 794 genes between the MPEF and control groups were found,361 genes of which were up-regulated while 433 were down-regulated.After MPEF treatment,caspase activity and pyruvate transport raised significantly whereas nucleotide metabolism,ribosomal biosynthesis,protein transcription and translation,RNA polymerase activity,superoxide dismutase activity,transmembrane transport regulation,vitamin metabolism were significantly down-regulated.Furthermore,6 differentially expressed genes were confirmed by real-time fluorescence quantitative polymerase chain reaction(q-PCR),achieving the same gene expression patterns as the transcriptome sequencing results.Destruction of cell structure,gene damage,restriction of protein synthesis and function,and changes in enzyme activities might be the main causes of death for Pichia after MPEF treatment.

关 键 词：毕赤酵母 MPEF杀菌技术 致死机理 转录组 实时荧光定量聚合酶链式反应 

分 类 号：TS201.3[轻工技术与工程—食品科学]