Project description:Hyperthermophilic archeaon, Thermococcus onnurineus NA1 has known as a strict anaerobe. To date, a few of studies have been reported that strict anaerobe can grow using oxygen. However, the research of the growth enhancement of strict anaerobic archaeon belonging to the order of Thermococcales using the oxygen, in which has never been reported so far. In this study, we showed that the growth of T. onnurineus NA1 strain increased under various oxygen concentrations and we observed that oxygen was decreased in the headspace during the growth of cell. Genome-wide transcriptomic analysis was carried out to evaluate alterations in gene expression induced by O2 and to explain the physiological effects of oxidative stress on the growth of T. onnurineus NA1.
Project description:Expression analysis of Thermococcus onnurineus NA1 KCTC10859 and an Frh-deficient mutant, Frh overexpression mutant and TON_0282 deletion mutant in a T. onnurineus NA1 strain.
Project description:A strong promoter increases transcription of the genes of interest and enhances the production of various valuable substances. For a hyperthermophilic archaeon Thermococcus onnurineus NA1, which can produce H2 by carbon monoxide oxidation, we searched for a novel endogenous strong promoter by transcriptome analysis using high-throughput RNA sequencing. Based on the relative transcript abundance, we selected one promoter to encode a hypothetical gene, of which homologs were found only in several Thermococcales strains. This promoter, PTN0510, was introduced into the front of CO-responsible hydrogenase gene cluster encoding a carbon monoxide dehydrogenase (CODH), a hydrogenase and a Na+/H+ antiporter. In the resulting mutant strain, KS0510, transcription and translation level of the gene cluster increased by 4- to 14-folds and 1.5- to 1.9-folds, respectively, in comparison with those of wild-type strain. Additionally, H2 production rate of KS0510 mutant was 4.8-fold higher than that of wild-type strain. The PTN0510 was identified to be much stronger than the well-known two strong promoters, gdh and slp promoters from Thermococcus strains, through RT-qPCR and western blotting analyses and kinetics of H2 production. In this study, we demonstrated that the RNA-seq approach is a good strategy to mine a novel strong promoter of use to a Thermococcus strain when developed as a biotechnologically promising strain to produce valuable metabolites through a metabolic engineering. RNA expreesion profile of T. onnurineus NA1 genes in CO-containing media, in duplicate, using Illumina HiSeq 2500
