|
|
Protein extraction from rice (Oryza sativa L.) root for two-dimensional electrophresis |
Xiaoliang XIANG1, Shuju NING2, Xia JIANG3, Xiaogui GONG1, Renlei ZHU1, Lanfang ZHU1, Daozhi WEI1() |
1. School of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou 350002, China; 2. School of Crop Sciences, Fujian Agriculture and Forestry University, Fuzhou 350002, China; 3. College of Chemistry and Materials Science, Fujian Normal University, Fuzhou 350002, China |
|
|
Abstract An efficient protein extraction method for two-dimensional gel electrophoresis (2-DE) from plant samples is usually challenging due to the low protein content and high level of interfering compounds. Proteomic analyses of rice (Oryza sativa L.) roots are limited by the lack of an efficient protein extraction method. To establish an effective protocol of protein extraction suitable for 2-DE analysis in rice roots, we evaluated three protein extraction methods (trichloroacetic acid [TCA]/acetone, Mg/NP-40/TCA, and tris-base/acetone). Our results showed that the Mg/NP-40/TCA extraction method had the highest protein yield and is the best resolution of protein separation among the three methods. The TCA/acetone method exhibited clear ?protein? profiles? and ?detected? more ?protein ?spots with the highest intensity in the region of high Mr (above 45 kDa) than the other methods. However, this method was unable to detect proteins with low-Mr (less than 24.0 kDa). The Tris-base/acetone method showed the poorest resolution of protein separation. Our results suggest that the Mg/NP-40/TCA method was the most effective among the three methods and may provide enhanced proteomic information for rice and other crop roots.
|
Keywords
protein extraction
two-dimensional electrophresis (2-DE)
rice (Oryza sativa L.) roots
|
Corresponding Author(s):
WEI Daozhi,Email:weidz888@sohu.com
|
Issue Date: 05 December 2010
|
|
1 |
Bradford M M (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem , 72(1-2): 248-254 doi: 10.1016/0003-2697(76)90527-3
|
2 |
Candiano G, Bruschi M, Musante L, Santucci L, Ghiggeri G M, Carnemolla B, Zardi L, Righetti P G (2004). Blue silver: A very sensitive colloidal Coomassie G-250 staining for proteome analysis. Electrophoresis , 25(9): 1327-1333 doi: 10.1002/elps.200305844
|
3 |
Damerval C, De Vienne D, Zivy M, Thiellement H (1986). Technical improvements in two-dimensional electrophoresis increase the level of genetic variation detected in wheat-seedling proteins. Electrophoresis , 7(1): 52-54 doi: 10.1002/elps.1150070108
|
4 |
G?rg A, Weiss W, Dunn M J (2004). Current two-dimensional electrophoresis technology for proteomics. Proteomics , 4(12): 3665-3685 doi: 10.1002/pmic.200401031
|
5 |
Hille J M, Freed A L, W?tzig H (2001). Possibilities to improve automation, speed and precision of proteome analysis: a comparison of two-dimensional electrophoresis and alternatives. Electrophoresis , 22(19): 4035-4052 doi: 10.1002/1522-2683(200111)22:19<4035::AID-ELPS4035>3.0.CO;2-9
|
6 |
Kim S T, Cho K S, Jang Y S, Kang K Y (2001). Two-dimensional electrophoretic analysis of rice proteins by polyethylene glycol fractionation for protein arrays. Electrophoresis , 22(10): 2103-2109 doi: 10.1002/1522-2683(200106)22:10<2103::AID-ELPS2103>3.0.CO;2-W
|
7 |
Laemmli U K (1970). Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature , 227(5259): 680-685 doi: 10.1038/227680a0
|
8 |
O’Farrell P H (1975). High resolution two-dimensional electrophoresis of proteins. J Biol Chem , 250(10): 4007-4021
|
9 |
Rabilloud T (1998). Use of thiourea to increase the solubility of membrane proteins in two-dimensional electrophoresis. Electrophoresis , 19(5): 758-760 doi: 10.1002/elps.1150190526
|
10 |
Shaw M M, Riederer B M (2003). Sample preparation for two-dimensional gel electrophoresis. Proteomics , 3(8): 1408-1417 doi: 10.1002/pmic.200300471
|
11 |
Xie C J, Wang D, Yang X Y (2009). Protein Extraction Methods Compatible with Proteomic Analysis for the Cotton Seedling. Crop Science Society of America , 49(2): 395-402
|
12 |
Xu C P, Xu Y, Huang B R (2008). Protein Extraction for Two-Dimensional Gel Electrophoresis of Proteomic Profiling in Turfgrass. Crop Science Society of America , 48(4): 1608-1614
|
13 |
Yang Q S, Wang Y Q, Zhang J J, Shi W P, Qian C M, Peng X X (2007). Identification of aluminum-responsive proteins in rice roots by a proteomic approach: cysteine synthase as a key player in Al response. Proteomics , 7(5): 737-749 doi: 10.1002/pmic.200600703
|
|
Viewed |
|
|
|
Full text
|
|
|
|
|
Abstract
|
|
|
|
|
Cited |
|
|
|
|
|
Shared |
|
|
|
|
|
Discussed |
|
|
|
|