Construction Of Alkaline Proteinase Gene Engineering Strains And Applied Research Of Spi Applied To Milk Beverages

Soybean protein isolate is a kind of vegetable protein which has a plenty of resources and high nutritivevalue. Soybean protein isolate was generally used to food produce with improving alimentary process functionand flavour through its unique function and characteristic. But the function and characteristics of Soybeanprotein isolate has some insufficient, the application was restrained. Enzymes modification is a important roadto improve soybean protein isolate function and characteristic. Through utilizing Bnfillus Licheniformis,technique for gene engineering was used to build alkaline proteinase, then the lkaline proteinase was uesd tomodification soybean protein isolate, and optimize technologcial conditions. Systematically to study andresearch the function of soybean protein isolate, which can be provided reference of milk beverages industry.The main research contents and conclusions are as follows:（1）The genomic DNA of Bacillus licheniformis for template, apr E gene sequence according to thedesign, synthesis apr primer, using the polymerase chain reaction (PCR) from Bacillus licheniformis DNAgenetic fragments in apr expansion, the cloning to intermediary carrier pGM-T sequenced in identification,will appraisal right plasmid enzyme cut on restructuring, connection, transformation, construction engineeringbacterium pET-22b-apr, and carries on the induction expression and identification, and determine the bestinduced time for4h, the best temperature for37℃induction. Folin method for the determination of the alkaliprotease enzymatic activity, enzyme activity for13000U· g-1.（2）By gene engineering bacterium to enzyme modified soybean protein isolated, through the singlefactor experiment and the orthogonal rotating combination optimization design, get the best protease modifiedsoybean protein isolated conditions for: the quality of soy protein concentration was6.86%, add proteaseconcentration for3120U/mL, enzyme solution effect time for64min; Through determinating the dissolubilityof soybean protein isolated of anterior-posterior modification, modified the solubility of soybean proteinisolated for94.01%, more before modification improved10.65%.（3）Through the comparative study of solubleness, emulsibility, viscosity and dispersity about SPI andESPI, the result shows that the ESPI has better solubleness, emulsibility, viscosity and dispersity, andfunctionality surpass SPI, which can be used in beverage industry. The ESPI is used adding to milk beverages,which can be substitute milk, and optimize the best ratable, get modified soybean protein isolated with milkratio for4:6, beverages with good sense and stable effect.