Method for extracting sucrose phosphate synthetase from muskmelon leaves and determining activity of sucrose phosphate synthetase

Abstract

The invention discloses a method for extracting sucrose phosphate synthetase from muskmelon leaves and determining activity of sucrose phosphate synthetase. The method comprises the following steps: (1), enzyme liquid extraction: taking fresh muskmelon leaves and putting the fresh muskmelon leaves into a mortar, grinding by an extraction buffer solution subjected to ice bathing, carrying out refrigerated centrifugation at a high speed after fully homogenizing, taking supernatant and desalting the supernatant by a centrifugal desalting column, wherein extracted enzyme liquid can be directly used for enzyme activity determination; and (2), enzyme activity determination: taking enzyme liquid, adding the enzyme liquid into a reaction substrate, wherein different reaction temperature and reaction time are determined and optimal reaction temperature and reaction time of sucrose phosphate synthetase in muskmelon leaves are determined, so that activity of enzyme can be reflected more accurately. According to the invention, salt, sugar, ions and other small molecular substances in enzyme liquid extracted primarily can be effectively removed, and reaction conditions of enzyme are optimized, so that the activity of sucrose phosphate synthetase in leaves can be determined more accurately.

Description

technical field [0001] The invention relates to a method for extracting sucrose phosphate synthase from melon leaves and measuring its activity. Background technique [0002] Sucrose phosphate synthase is a key enzyme of sucrose metabolism and an important photosynthetic enzyme. Its activity is positively correlated with the net photosynthetic rate. Therefore, the study of this enzyme activity is of great significance for improving the photosynthesis of melon. At present, G-25Sephadex column and dialysis bag are mostly used for the extraction of sucrose phosphate synthase. The G-25Sephadex column is expensive and the extraction cost is high, while the price of dialysis bag is low, but the operation is complicated, time-consuming, and the extraction efficiency is low; Centrifugal desalting columns are slightly more expensive, but they are reproducible, and their extraction efficiency is better than that of dialysis bags and G-25Sephadex columns. In terms of enzyme activity m...

AI Technical Summary

Problems solved by technology

At present, G-25Sephadex column and dialysis bag are mostly used for the extraction of sucrose phosphate synthase. The G-25Sephadex column is expensive and the extraction cost is high, while the price of dialysis bag is low, but the operation is complicated, time-consuming, and the extraction efficiency is low; Centrifugal desalting columns are slightly more expensive, but they are reproducible and have better extraction efficiency than dialysis bags and G-25Sephadex columns

In terms of enzyme activity determination, the current problem is that researchers pay less attention to the reaction time and temperature of enzyme and substrate, and different materials have different optimal reaction conditions. The present invention optimizes the conditions for the determination of sucrose phosphate synthase activity in melon leaves

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