Study On The Detection Method Of The Phytase Activity In Transgenic Corn

Phytase is general term of the enzymes which catalyze hydrolysis of phytic acid and phytate into cyclohexanehexol and phosphoric acid (or phosphate), belongs to monoester phosphohydrolase. Phytase can break down phytic acid to release a large number of inorganic phosphorus.If put the phytase in animal feed, monogastric animals can break down large amounts of phytic acid into phosphorus, then the organic phosphorus absorption rate increased, and the discharge of phosphorus reduced, the environmental pollution also reduced. There are relevant national standards on methods for measuring phytase activity which is added into fodder, but reports on methods of measuring phytase activity in plants are less, therefore study on the detection method of the phytase activity extractived from plants has practical significance.Objectives:To determine the best method applied for phytase extraction and activity measurement in transgenic corn through optimizing the condition of phytase extraction and measurement, and monitoring the phytase activity of the transgenic corn grows in the field and in the pot at different growth stages. To provide evidence for evaluating phytase transgenic corn.Method:The univariates related with phytase extraction such as extraction time, the ratio of sample to extract and the pH of extracting solution were analyzed and the level of experimental factors with higher phytase activity is selected.Then optimize extraction conditions according to the L9(33) orthogonal layout.The univariates related with phytase measure such as reaction time.zymolyte concentration.the pH of buffer were analyzed and the level of experimental factors with higher phytase activity is selected.Then optimize measuring conditions according to the L9(33) orthogonal layout.The phytase activity of corn grows in the field and in the pot at different growth stages were detected by the method of our studied.Results:The result of the phytase extraction experiments shows that when the ratio of sample to extract is 1:4, the extraction time is 15min, pH is 5.50, the phytase activity and extraction rate was highest.The result of the measuring methods of the phytase activity shows that when the reaction time was 30min. buffer pH is 5.50,the zymolyte concentration was 0.1Ommol/L. phytase activity is the highest.Compared with control group, phytase activity in leaves. stems and roots of transgenic corn grows in the field and pot at different growth stages is higher. but there were no significant difference. but phytase activity in the seeds is higher.Conclusion:The conclusions according to the results of this study were as follows:1. The best condition of phytase extraction from the transgenic corn by ultrasonic method are that ratio of sample to extract is 1:4. the extraction time is 15min. the pH of extracting solution is 5.50.2. The best condition of phytase measurement using spectrophotometry are that the reaction time is 30min. the pH of buffer is 5.50.and the zymolyte concentration was 0.10mmol/L3. The phytase activity in the leaves, stems and roots of the transgenic corn grows in the field and pot is higher than control group, and phytase activity in seeds is much higher than control group. Keywords: Phytase. Transgenic Corn. Detection Method. Orthogonal Test...