[目的]基于转录组学技术研究表达磷脂酶A2的毕赤酵母重组菌在甲醇诱导表达外源蛋白时的基因表达差异,从而解析外源蛋白高效诱导表达机制,为进一步工程菌株的改造提供理论支撑.[方法]以一株产磷脂酶(PLA2)的毕赤酵母为出发菌株,采用RNA-Seq二代测序方法,研究在甘油培养和甲醇诱导两种条件下,重组毕赤酵母转录组基因表达差异情况.[结果]重组毕赤酵母中共鉴定到5225个转录本.甘油培养与甲醇诱导相比,共有857个基因发生显著变化.依据代谢途径分类,差异基因集中在核糖体成分、甲醇代谢、磷酸戊糖途径、糖酵解途径、柠檬酸循环、乙醛酸循环以及蛋白质加工过程.[结论]通过分析甲醇诱导前后的差异表达基因,结果表明碳源改变对胞内代谢会产生全局影响.本研究结果为进一步研究毕赤酵母表达外源蛋白的机制提供了基础.%[Objectives] In order to study the mechanism of heterologous protein expression in Komagataella phaffii,transcriptomics technology was used to study the profile of differentially expressed genes in the recombinant K.phaffii during the methanol-induced expression of phospholipase A2.[Methods] We used RNA sequencing (RNA-seq) technology to analyze the differentially expressed genes in the recombinant K.phaffii expressing phospholipase A2 between glycerol and methanol cultivation condition.[Results] In total 5225 transcripts were identified in the recombinant K.phaffii.Compared between glycerol and methanol cultivation condition,857 genes were significant differentially expressed.According to KEGG (Kyoto Encyclopedia of Genes and Genomes) analysis,the differentially expressed genes were mostly related to ribosome composition,methanol metabolism,pentose phosphate pathway,citric acid cycle,glyoxylic acid cycle and protein processing process.[Conclusions] A global effect on the metabolism was observed when changing the carbon source from glycerol to methanol for the induction of heterologous protein.Our results provide rich information for further in-depth studies of the mechanism of protein expression in K.phaffii.
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