基于转录组学挖掘兽疫链球菌内源表达元件及高产透明 质酸应用.

【Objective】This work is aimed at establishing a library of endogenous expression elements of Streptococcus epizooticus, and investigating the application of it in increasing hyaluronic acid yields.【Method】The 32 kinds of high, medium and low strength promoter and RBS combination (expression element, PR) were preliminarily screened, through transcriptomics analysis of S. zooepidemicus in the exponential or stationary stage of fermentation. Then, the strength of the 32 PRs was verified via detecting the transcriptional level of gfp gene and fluorescence intensity of GFP. Eventually, the key hyaluronic acid synthesis gene hasA, hasB, hasC, hasD, and hasE was overexpressed respectively with the selected strong expression element PR31, and the impact of the strong PR on enhancing hyaluronic acid production was assessed through a 2 L fermentation.【Result】The relative transcription of the aforementioned genes were elevated to 8.17, 7.32, 3.72, 39.48 and 9-fold. Among them, the overexpression of hasA and hasD increased the hyaluronic acid yield by 43% and 31%, respectively, compared to the wild type, reaching 5.654 g/L and 5.283 g/L. 【Conclusion】A library of endogenous expression elements of S. zooepidemicus had been constructed successfully, which can be employed for metabolic engineering, such as reinforcing the hyaluronic acid synthesis pathway or weaking competitive pathways. [ABSTRACT FROM AUTHOR]