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Published on: March 19, 2015
[Establishment of two-dimensional electrophoresis system of caudal gland]
Shu-fen Li1, Feng Wang, Gao-hui Zhang
1Institute of Special Wild Economic Animal and Plant Science, Chinese Academy of Agricultural Sciences, Changchun 132109, China. lishufen1122@163.com
Objective:
To establish an effective separation system of 2-DE for the proteome of caudal gland, and provide foundation for revealing the mechanisms of histological development and pharmacological activities.
Method:
The total proteins of caudal gland were extracted by TCA/acetone precipitation, phenol extraction/methanol-ammonium acetate precipitation and trizol-base method respectively and separated by immobilized pH gradient (IPG) strips prior to SDS-PAGE. Loading protein sample size and isoelectric focusing conditions were optimized. The gels were stained with Coomassie brilliant blue, scanned and then analyzed using PDQuest 8.0 analysis software.
Result:
The total proteins of caudal gland extracted by trizol-base method were the highest quality and could meet the needs of 2-DE. With 300 microg of proteins loaded on 7 cm pH 3-10 IPG strip followed by isoelectric focusing program II ,a satisfying 2-DE profiles were obtained. The total number of disticted protein spots was 209 with the optimized system.
Conclusion:
A well-resolved 2-DE patterns of caudal gland were obtained by this optimized system. This method could be applied to prepare other similar tissue sample and 2-DE studies.
