The Preliminary Bioinformatics Analysis Of A Novel TEH₀5770 Gene From Tetragenococcus Halophilus And Its Heterologous Expression

Tetragenococcus halophilus,isolated from high salt liquid soy mash in the traditional soy sauce fermentation process,could grow at an upper limit of 3.5 M,with a crucial role in flavor formation of high salt food fermentation due to its wide growth NaCl concentration range.Recent researches mainly focus on molecular chaperone and compatible solutes to study the salt-tolerant mechanism of T.halophilus,however,only limited to some known genes,dnaK,clpB,some known compatible solutes related genes like butA et al.However,research showed there were still some unknown function genes involved in T.halophilus salt tolerance mechanism.Here we mainly study one novel gene in T.halophilus,TEH₀5770's expression level under salt stress conditions.In this study,we would do some preliminary bioinformatics analysis of the novel gene TEH₀5770 to preliminary define the physical and chemical propertiesof related protein,and then we used molecular biological techniques to analyze the expression levels of TEH₀5770 in different salt concentration media,both RNA and relatied protein's expresstion level;and we also built a heterologous expression vector and transformed it to Escherichia coli BL21,and used SDS-PAGE and western blot to measure the expression level in recombinant E.coli to figure out the expression level of TEH₀5770 gene on the recombinant BL21.In addition,we also measured the growth curve assay of recombined E.coli to preliminary define whether the novel gene TEH₀5770's transcriptional and expression level were influenced by salt concentration in the external environment.RT-PCR results showed that TEH₀5770 gene transcriptional level was up-regulated under salt stress conditions,and so did the protein expression level in T.halophilus.SDS-PAGE and western blot analysis indicated successful expression of recombinant TEH₀5770 gene in E.coli strain BL21.Recombinant E.coli containing TEH₀5770 gene exhibited an obvious growth advantage under high salt concentration,which revealed that TEH₀5770 gene of T.halophilus could respond to external environmental salt concentration indeed.Altogether,the bioinformatics analysis of the novel gene TEH₀5770 and researches on expression level under different salt concentration in T.halophilus,and in heterologous hosts preliminary defined that the novel gene's transcriptional and expression level were influenced by salt concentration.These researches laid a foundation for researching TEH₀5770 gene's function and involved metabolic pathways in our laboratory.