Chinese Bulletin of Botany ›› 2015, Vol. 50 ›› Issue (6): 739-745.DOI: 10.11983/CBB14186

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Extraction and Separation Analysis of Total Protein from the Bark of Apple Branches

Caixia Zhang1,2, Yi Tian1,2, Liyi Zhang1,2, Long Xiao1,2, Peihua Cong1,2*   

  1. 1Key Laboratory of Horticulture Crops Germplasm Resources Utilization, Ministry of Agriculture, Research Institute of Pomology, Chinese Academy of Agricultural Sciences, Xingcheng 125100, China
    2Research Institute of Pomology, Chinese Academy of Agricultural Sciences, Xingcheng 125100, China
  • Received:2014-10-27 Accepted:2015-01-27 Online:2015-11-01 Published:2015-09-06
  • Contact: Cong Peihua
  • About author:

    ? These authors contributed equally to this paper

Abstract:

We aimed to establish a suitable extraction method for analyzing total protein in the bark of apple branches. The branches of an 8-year-old cultivar (Huayue) were used as material. We compared 3 different extraction methods, including TCA-acetone precipitation (A), methanol/ammonium acetate precipitation (B), and improved Tris-phenol extraction (C). We optimized the extraction procedures and established an ideal method for analyzing the total protein in the bark of apple branches. The improved Tris-phenol extraction method could reveal a good map of 993 protein spots, significantly more than the other two methods: 418 and 674 protein spots were obtained with TCA-acetone precipitation and methanol/ammonium acetate precipitation, respectively. Furthermore, the gel background was clear, and the protein spots were better focused than with the two other methods. After comparing sample loading quantity, we selected 800 μg as the ideal loading quantity for our study. Furthermore, we selected several proteins for MALDI-TOF-TOF/MS analysis and database searching, and the protein spots were finally identified. In this study, we optimized the extraction procedures and sample loading quantity for total protein and finally obtained an ideal 2-D electrophoresis map. Our study may provide a good basis for further proteome analysis of apple branches.