Selenium-induced Expression Of Selenium-containing ZEIN Protein By Bacillus Subtilis

Selenium?Se?is an essential trace element in organisms and plays an important role in antioxidation,prevention of cancer,and enhancement of immunity.Met is an essential amino acid for organisms and participates in the synthesis of proteins and affects the growth and development of the organism;neither of them can be synthesized by auto metabolism,must be taken from the body.In the body,Se binds to Met to form Se-Met.Se-Met has the functions of promoting the growth of the body,enhancing immunity,and inhibiting tumor growth.In view of the current situation of selenium deficiency in most parts of China,this study was to establish and express zein with high Met content on the basis of screening and identification of Bacillus subtilis-resistant bacteria,and to obtain selenium-rich Bacillus subtilis and its expression.The product selenium-containing zein?Se-ZEIN?provides a new way for humans and animals to supply Se-Met.Using different concentrations of Na2SeO3 as screening conditions,the differences in selenium-growth capacity of wild-type,168,WB600,WB800,and WB800N Bacillus subtilis strains were obtained,and WB800N strains with good growth conditions with Na2SeO3 concentration as high as 20 mM were determined as the host bacteria of the subsequent experiments.The zein gene?NCBI gene accession number:541922 dzs10,fragment length of 486 bp?was obtained by chemical synthesis and was linked to E.coli Bacillus subtilis shuttle vector p HT43.Recombinant plasmid pHT43-zein;recombinant plasmid pHT43-zein was transformed into Bacillus subtilis WB800N to obtain the recombinant strain WB800N-pHT43-zein,and WB800N-pHT43-zein was cultured in Na₂SeO₃ medium to obtain Se with a molecular weight of approximately 18 kDa.The residual amount of Na₂SeO₃ and the total selenium content of the bacteria in the supernatant of the culture supernatant were measured by methylene blue and DTNB methods,respectively,and the selenium conversion rate of the recombinant bacteria was 74.58%.In the single factor experiment,the conventional culture condition of the recombinant bacteria was 37°C,180rpm,pH 7.0 and inoculation amount 6%.After optimizing the culture conditions by response surface method,the selenium conversion rate was increased to 89.77%.The number of selenium combined with Se-ZEIN reaches5.2.