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

Abstract: 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 high-throughput 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.

Key words: Pichia, MPEF sterilization, lethal mechanism, transcriptome, real-time fluorescence quantitative polymerase chain reaction (q-PCR)