Optional Protein Extracting Methods of Poplar Leaves for Two-dimensional Gel Electrophoresis

doi:10.7525/j.issn.1673-5102.2016.03.022

Abstract

Abstract: In order to obtain the two-dimensional gel electrophoresis images with better separation and definition of poplar leaves, the Populus X, P.trichocarpa Torr. & Gray and P.alba×P.glandulosa were selected to study the optional extracting methods of poplar leaves for two-dimensional gel electrophoresis by using TCA/acetone precipitation, Tris/saturated phenol and Tris/TCA method, respectively. The proteins of poplar leaves were separated by SDS-polyacrylamide gel electrophoresis and two-dimensional gel electrophoresis respectively and the images of two-dimensional gel electrophoresis were analyzed by Image Master 2D Platinum 6.0. Tris/TCA method was more appropriate for the protein extraction of poplar leaves for two-dimensional gel electrophoresis. The average protein content of the fresh sample extracted by Tris/TCA method was 949.46 μg·g^-1 and the average quantity of protein spots was 567 in the two-dimensional gel electrophoresis images which had better separation and definition.

Key words: poplar leaves, protein extraction, Tris/TCA method, two-dimensional gel electrophoresis

CLC Number:

S792.11

YAN Dong, LIU Dian-Kun, SI Dong-Jing, ZHENG Mi, ZHAO Xi-Yang, QU Guan-Zheng, LI Ying. Optional Protein Extracting Methods of Poplar Leaves for Two-dimensional Gel Electrophoresis[J]. Bulletin of Botanical Research, 2016, 36(3): 469-475.

References

1. O'Farrell P H.High resolution two-dimensional electrophoresis of proteins[J].J Biol Chem,1975,250(10):4007- 4021.
2. Komatsu S,Konishi H.Proteome analysis of rice root proteins regulated by gibberellin[J].Genomics,Proteomics & Bioinformatics,2005,3(3):132-142.
3. Nozu Y,Tsugita A,Kamijo K.Proteomic analysis of rice leaf,stem and root tissues during growth course[J].Proteomics,2006,6(12):3665-3670.
4. Chen F,Yuan Y X,Li Q,et al.Proteomic analysis of rice plasma membrane reveals proteins involved in early defense response to bacterial blight[J].Proteomics,2007,7(9):1529-1539.
5. Manaa A,Ahmed H B,Valot B,et al.Salt and genotype impact on plant physiology and root proteome variations in tomato[J].Exp Bot,2011,62(8):2797-2813.
6. Chen Q,Zhang M D,Shen S H.Comparison of protein extraction methods suitable for protemics analysis in seedling roots of Jerusalem artichoke under salt(NaCl) stress[J].Afr.J.Biotechnol.,2011,10(9):7650-7657.
7. Damerval C,Vienne D D,Zivy M,et al.Technical improvements in two-dimensional electrophoresis increase the level of genetic variation detected in wheat seedling proteins[J].Electrophoresis,1986,7(1):52-54.
8. 刘海衡,胡胜武,刘胜毅,等.油菜不同器官高质量总蛋白提取方法和双向电泳体系的优化[J].中国油料作物学报,2009,4(4):426-433. Liu H H,Hu S W,Liu S Y,et al.Optimization of high quality total protein extraction and two-dimensional gel electrophoresis system for different Brassica napus organs[J].Chinese journal of oil crop sciences,2009,4(4):426-433.
9. Saravanan R S,Rose J K C.A critical evaluation of sample extraction techniques for enhanced proteomic analysis of recalcitrant plant tissues[J].Proteomics,2004,4(9):2522-2532.
10. Carpentier S C,Witters E,Laukens K.Preparation of protein extracts from recalcitrant plant tissues:An evaluation of different methods for two dimensional gel electrophoresis analysis[J].Proteomics,2005,5(10):2497-2507.
11. 张颖,王猛,柴娜.杨树叶片离体再生系统的构建[J].山东农业科学,2010,5:8-10. Zhang Y,Wang M,Chai N.Establishment of in vitro regeneration system of poplar leaves[J].Shangdong Agricultural Sciences,2010,5:8-10.
12. Bradshaw HD,Ceulemans R,Davis J,et al.Emerging model system in plant biology:poplar(Populus) as a model forest tree[J].Journal of Plant Growth Regulation,2000,19(3):306-313.
13. Taylor G.Populus:Arabidopsis for forestry.Do we need a model tree?[J].Ann.Bot.2002,90(6):681-689.
14. Tuskan G A,DiFazio S,Jansson S,et al.The genome of black cottonwood,Populus trichocarpa(Torr.&Gray)[J].Science,2006,313(5793):1596-1604.
15. Tuskan G A,DiFazio S ,Teichmann T.Poplar genomics is getting poplar:the impact of the poplar genome project on tree research[J].Plant Biology,2004,6(1):2-4.
16. Guo Y M,Shen S H,Jing Y X,et al.Plant proteomics in the post-genomicera[J].Act a Bot Sin,2002,44(6):631-641.
17. Ramagli L S,Link A J.Methods in molecular biology[M].Totowa:Humana Press,1999.
18. 时立文.SPSS 19.0统计分析从入门到精通[M].清华大学出版社,2012:28-29. Shi LW.SPSS 19.0 statistical analysis from entry to the master[M].Tsinghua university press,2012:28-29.
19. 李晓琳,邵爱娟,陈敏,等.酸浆种子蛋白提取方法的比较研究[J].中国实验方剂学杂志,2010,16(1):14-17. Li X L,Shao A J,Chen M,et al.Comparison of protein extraction methods for the semen seu sructus shysalis[J].Chinese Journal of Experimental Traditional Medical Formulae,2010,16(1):14-17.
20. 王卓伟,余茂德,鲁成.PVP在桑叶总DNA提取中的应用[J].西南大学学报:自然科学版,2001,23(1):61-62. Wang Z W,Yu M D,Lu C.Application of PVP in total DNA extraction in mulberry[J].Journal of Southwest Agricultural University(Natural Science Edition),2001,23(1):61-62.
21. Muhammad A Lodhi,Guang-Ning Ye,Norman F Weeden,et al.A simple and efficient method for DNA extraction from grapevine cultivars and Vitis species[J].Plant Molecular Biology Reporter,1994,12(1):6-13.
22. Islam N,Lonsdale M,Upadhyaya N M,et al.Protein extraction from mature rice leaves for two-dimensional gel electrophoresis and its application in proteome analysis[J].Proteomics,2004,4(7):1903-1908.
23. 赵相涛,吕全,赵嘉平,等.一种适合杨树树皮的蛋白质提取方法[J].山东农业科学,2012,44(7):121-123. Zhao X T,Lü Q,Zhao J P,et al.A protein extraction method suitable for poplar bark[J].Shangdong Agricultural Sciences,2012,44(7):121-123.
24. 袁坤,王明庥,黄敏仁.一种适合杨树叶片的蛋白质提取方法[J].南京林业大学学报:自然科学版,2007,31(3):119-121. Yuan K,Wang M X,Huang M R.A method suitable for protein extraction of poplar leaves[J].Journal of Nanjing Forestry University(Natural Science Edition),2007,31(3):119-121.