| Polyvinyl alcohol(PVA),as a kind of high performance polymer materials,was used in many fields of production and every aspect of science research.It has become one of the largest demanded polymer materials of the world.The microbial isolation and application research on reducing the polyvinyl alcohol pollution to the environment is of great importance.This research revolves around a strain HK1 which grows with polyvinyl alcohol as the only carbon source.The research includes identification of strain,optimizing the growth conditions of strain,whole-genome sequencing,gene knockout,and expressing the degradation gene in E.coli.The main results of these points were as followed.(1)Strain HK1 colony in LB medium presents an irregular shape,with light yellow color,opaque,surface drying with fold,uplift in the colony central and rim jagged.The strain HK1 was characterized as Bacillus amyloliquefaciens by testing its physiology,biochemistry,and molecular identification.(2)Through the lethal concentration detection of HK1 strains with different antibiotics,it can be seen that strain HK1 is resistant to Tetracycline,chloramphenicol,and spectinomycin hydrochloride.To streptomycin,gentamycin,kanamycin,erythromycin,penicillin and ampicillin,strain HK1 has no resistance.(3)The optimal conditions for PVA biodegradation of the obtained strain were30?,1.0g/LPVA,p H8.0,and 2.0 g/L yeast.(4)Selected a DNA sequence from the whole genome sequencing information successfully,which was predicted a quinone structure protein ethanol dehydrogenase domain of membrane protein gene named 3057.Gene 3057 has been linked successfully to plasmid p ET-28 a to constructe an expression vector,and the expression vector was transformed into E.coli BL21.In addition,3057 gene has been successfully expressed by Western Blot test.Through the thick enzyme fluid experiment,it can be proved that 3057 gene in vitro expression product has the activity of degradation of polyvinyl alcohol. |
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