Screening And Characteristics Of Selenium-Enriched Yeast Isolated From Mulberry Orchard Of Kaiyang

Selenium is an essential trace element for human body with various physiological functions,and the selenium intake of most people in China is insufficient.Therefore,the research and development of selenium-enriched products is of great significance for improving the national health level.As an organic selenium source,selenium-enriched yeast has higher biological activity and safety than inorganic selenium source.It is an ideal additive for selenium-enriched functional products and has broad application prospects.In this study,the mulberry orchard in the selenium-rich region of Kaiyang,Guizhou was used as the source of yeast isolation.A yeast with strong selenium-enriching ability was selected by qualitative and quantitative methods,and molecular biology identification and growth characteristics were explored.At the same time,the selenium-enriched fermentation conditions of the strain were optimized,the amino acid composition of the selenium-enriched yeast was analyzed,and finally the selenium-enriched mechanism of the yeast was preliminarily explored based on transcriptomics technology,with a view to further development of the selenium-enriched yeast use for reference.The main results of the study are as follows:?1?A total of 75 strains of yeast were isolated from a mulberry orchard in Kaiyang,a selenium-rich area,and then selenium-tolerant method and red selenium method were used for initial selecting,and then selenium conversion rate was used for rescreening to obtain a selenium conversion.The yeast with a higher rate was numbered as SY2,which has a selenium-enriched amount of 2278.35?g/g,a biomass of 4.31 g/L and a selenium conversion rate of 49.11%.The initial identification of WL medium and 26S r DNA sequence analysis identified the strain is Saccharomyces cerevisiae.?2?Study the growth characteristics of S.cerevisiae SY2.The lag period was0?2 h,and the logarithmic period was 2?10 h.After that,it entered a stable period.It has the characteristics of large gas production,its optimal growth temperature is28?,optimal fermentation temperature is 30?,lethal temperature is 68?,optimal growth p H is 4.5,optimal fermentation p H is 4.5,and SO₂concentration is 260mg/100 m L,alcohol-resistant concentration of 14%?v/v?,and the basic fermentation medium is the best selenium-rich medium.The Box-Behnken central combination experiment was used to determine the optimal selenium-enriched culture conditions:selenium addition time was 6 h after cultivation,culture temperature was 30?,initial p H was 4.5,inoculation amount was 6%,selenium concentration was 20.00?g/m L,the liquid volume was 100/250 m L,and the culture was carried out on a 150 r/min thermostatic shaker for 72 h.Under this condition,the biomass was 5.40 g/L,the selenium content was 2427.24?g/g,and the conversion rate was 65.50%.The results of amino acid analysis of yeasts before and after selenium enrichment showed that the total amino acid content of yeasts without selenium enrichment was 418.57 mg/g,the essential amino acid content was 146.56 mg/g,the total amino acid content of yeast after selenium enrichment was 431.83 mg/g,the content of essential amino acids is175.77 mg/g,they have increased by 3.17%and 19.93%,respectively.In addition,the Met is also added to the selenium-enriched yeast,indicating that the selenium-enriched yeast has a higher nutritional value.?3?The Illumina Novaseq 6000 platform was used for high-throughput sequencing of the two yeast strains before and after selenium enrichment,and analysis was performed by transcriptomics analysis.The results showed that 994 differentially expressed genes were identified,of which 498 genes are down-regulated;496 genes are up-regulated.KEGG Pathway enrichment analysis found that differentially expressed genes were successfully annotated into 113 KEGG metabolic pathways,mainly including“meiosis-yeast”,“glycolysis/gluconeogenesis”,“sulfur metabolism”,“cysteine acid and methionine metabolism”,“Glutathione metabolism”and“Selenium compound metabolism”etc.The 9 differentially expressed genes obtained by analysis are named by MET22,MET5,SPE4,MEU1,GSH2,ZWF1,STR2,TRR1,YML082W,They mainly encode 3'?2'?,5'-bisphosphate nuclease and subunit?-sulfite reductase related to sulfur metabolism;spermine synthase and S-methyl-5-thioadenos-ine phosphorylase related to cysteine and methionine metabolism;glutathione synthase and glucose 6-phosphate dehydrogenase related to glutathione metabolism and cystathionine?synthase,thioredoxin-disulfide bond reductase and putative cystathionine?synthase,which are involved in the metabolism of selenium compounds,and they all showed up.Based on comprehensive analysis,it is speculated that the enrichment of selenium by strain S.cerevisiae SY2 is mainly that in the selenium-rich state,selenium enters the cell through the sulfate transporter and reacts with glutathione to form selenium.Glutathione?GS-Se-SG?is then reduced to hydrogen selenide by the reducing agent in the cell,which is further converted into selenoamino acids and selenoproteins.At the same time,many genes involved in metabolic pathways are also activated or increased,which further promotes the synthesis and intracellular enrichment of organic selenium.