水稻可溶性糖蛋白的纯化与鉴定研究

从水稻鲜叶中提取总蛋白，对总蛋白中的蛋白质含量进行了测定；通过硫酸铵沉淀将总蛋白提取液进行分级，从而达到了总蛋白细分和放量的目的。四级份的分级蛋白分别通过ConA-Sepharose 4B 亲和层析进行糖蛋白纯化，按吸收峰收集的各级糖蛋白混合物进行冷冻干燥，得到干粉；结合PAS法染色和考马斯亮蓝R-250染色对四级份的糖蛋白样品鉴定，在其中3个级份中均检测出糖蛋白；由于感度的差异，按考马斯亮蓝R-250染色可检测出近30种糖蛋白(包括部分糖肽)，按PAS法染色可检测到7种糖蛋白；对3种含量较高的糖蛋白进行了胶上纯化，3种糖蛋白的PAS法染色均证实了3种样品为单一的糖蛋白或者糖肽，分别命名为RG1、RG2和RG3。

Purification and identification of rice soluble glycoproteins

The total proteins was extracted from rice leaves,and quantified by Coomassie Brilliant Blue G-250.We got four classifications from rice total proteins by using Ammonium Sulfate Precipitation,which was very important because we got easy way to separate and classify the total proteins.Also,it was useful for the purification of glycoproteins.The four classifications then purified by using ConA-Sepharose 4B Affinity Chromatography,the liquid of glycoproteins was collected by tubes in the same time of the appearances of absorption peaks.We even got some dry samples by using vacuum freezing and drying technology.Next,we done SDS-PAGE and determination respectively by Periodic Acid Schiff(PAS) staining and Coomassie Brilliant Blue(CBB) R-250 staining.In the gel of CBB staining,there are near 30 strips of different glycoproteins(some are glycopeptides) in the last three classifications;but the gel of PAS staining only showed 7 strips of different glycoproteins(some are glycopeptides) only because of the sensitivities of PAS much lower than CBB staining.The last step,we choose three glycoproteins showed on the gel of 50%~80% glycoproteins classification,all of them have higher contents than others.We further purified each of the three glycoproteins directly from the SDS-PAGE gels.They are proved purified glycoproteins or glycopeptides by CBB and PAS staining again.The three glycoproteins are named respectively as RG1,RG2 and RG3.