勃氏甜龙竹笋蛋白的提取及双向电泳分析
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摘要
为建立一套适于勃氏甜龙竹笋蛋白的双向电泳体系,分别采用TCA/丙酮法和改良TCA-酚法提取竹笋总蛋白,pH 3～10和pH 4～7的IPG胶条进行蛋白双向电泳研究。结果表明,TCA/丙酮法更适合勃氏甜龙笋总蛋白的提取,不仅具有更高的提取率,而且还能提取更多的蛋白种类;采用24 cm,pH 4～7线性IPG胶条,共分离出约1 196个蛋白点,比pH 3～10胶条获得更多的蛋白点数量,且蛋白点清晰可见,分辨率更高,分离效果更好。勃氏甜龙竹的竹笋蛋白主要是以等电点在pH 4～7,分子量集中在10～40 kDa范围内的低分子量酸性蛋白为主。
In order to establish a suitable two-dimensional gel electrophoresis system for the protein extraction of Dendrocalamus brandisi bamboo shoots,the total protein of bamboo shoots was extracted by TCA/acetone and modified TCA-phenol methods respectively,and the two-dimensional gel electrophoresis was performed on IPG strips with pH 3-10 and pH 4-7. The results showed that the TCA/acetone method was more suitable for the total protein extraction of Dendrocalamus brandisi bamboo shoots,obtaining a higher extraction rate and more types of protein. Using 24 cm pH 4-7 linear IPG strip,a total of 1,196 protein spots were isolated,much more than that by using pH 3-10 strip,and the protein spots were clearly visible,with higher resolution and better separation effect. The protein of Dendrocalamus brandisi bamboo shoots is mainly low molecular weight acidic protein with isoelectric point at pH 4-7 and its molecular weight concentrated in the range of 10-40 kDa.
In order to establish a suitable two-dimensional gel electrophoresis system for the protein extraction of Dendrocalamus brandisi bamboo shoots,the total protein of bamboo shoots was extracted by TCA/acetone and modified TCA-phenol methods respectively,and the two-dimensional gel electrophoresis was performed on IPG strips with pH 3-10 and pH 4-7. The results showed that the TCA/acetone method was more suitable for the total protein extraction of Dendrocalamus brandisi bamboo shoots,obtaining a higher extraction rate and more types of protein. Using 24 cm pH 4-7 linear IPG strip,a total of 1,196 protein spots were isolated,much more than that by using pH 3-10 strip,and the protein spots were clearly visible,with higher resolution and better separation effect. The protein of Dendrocalamus brandisi bamboo shoots is mainly low molecular weight acidic protein with isoelectric point at pH 4-7 and its molecular weight concentrated in the range of 10-40 kDa.
引文
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[2] 杨宇明,辉朝茂.优质笋用竹产业化开发[M].北京:中国林业出版社.1998:84.
[3] 裴佳龙,李鹏程,王茜,等.云南不同地理种源勃氏甜龙竹竹笋营养成分比较[J].西北林学院学报,2018,33(1):156-161.
[4] Koh J,Chen S X,Zhu N,et al.Comparative proteomics of the recently and recurrently formed natural allopolyploid Tragopogon mirus (Asteraceae) and its parents,New Phytologist,2012,196(1):292-305.
[5] 罗泽宇,杨粤军,刘选明.拟南芥蛋白质组研究中双向电泳技术条件的优化[J].激光生物学报,2008,17(4):539-543.
[6] 杜文鹏,徐彭,刘波,等.毛竹笋化学成分研究(Ⅰ)[J].中草药,2015,46(3):334-338.
[7] 李斌,张勇,杜文鹏,等.毛竹笋化学成分研究(Ⅱ)[J].中药材,2015,38(12):2535-2537.
[8] 朱飞冀,詹儒林,柳凤,等.芒果叶片双向电泳体系的建立及优化[J].基因组学与应用生物学,2017,36(2):814-819.
[9] Ong S E,Pandey A.An evaluation of the use of two-dimensional gel electrophoresis in proteomics[J].Biomolecular Engineering,2001,18(5):195-205.
[10] Dai S J,Li L,Chen T T,et al.Proteomic analyses of Oryza sativamature pollen reveal vel proteins associated with pollen germination and tube growth[J].Proteomics,2006,6(8):2504-2529.
[11] Dai S J,Chong K,Xue Y,et al.Proteomics identification of differentially expressed proteins associated with pollen germination and tube growth reveals characteristics of germinated Oryza sativa pollen[J].Molecular Cellular Proteomics,2007,6:207-230.
[12] 王海燕,田维敏,王斌,等.巴西橡胶树幼叶和成熟叶比较蛋白质组学初步研究[J].中国农学通报,2012,28(25):6-14.
[13] 杨慧敏,吴良如,潘雁红,等.毛竹笋蛋白的双向电泳分析[J].竹子研究汇刊,2015,34(3):35-38
[14] 潘雁红,杨慧敏,吴良如,等.雷竹笋蛋白的双向电泳分析[J].竹子学报,2017,36(1):36-40.