title: A novel method of protein extraction from perennial Bupleurum root for 2-DE.
abstract: The perennial Bupleurum root is thick and woody and contains high levels of interfering compounds. Common protein extraction methods have proved refractory towards the isolation of proteins suitable for 2-DE, due to the presence of interfering compounds. A novel method for extracting proteins suitable for 2-DE was established to overcome these problems. The main characteristic of this protocol is the partitioning of the proteins into the aqueous (fraction A-2), chloroform and isoamyl alcohol phases (A-3), and the interphase (A-1). The proteins are then extracted from each of these phases. From A-1, 85% (extracted protein against total proteins) proteins could be extracted and purified. For fraction A-2, a novel phenol extraction step is employed for the extraction of proteins. Based on the well-resolved 2-DE patterns, our protein preparation is free of interfering compounds. Using these methods (A-1, A-2, and A-3-3), a total of 3662 (1526 + 1128 + 1008) spots could be separated, and a protein yield of about 1.41 mg per 1.0 g fresh root material was obtained. To our knowledge, this is the first time that a protocol for protein extraction from perennial Bupleurum root has been reported that gives reproducible results. The protocol is expected to be applicable to other recalcitrant plant tissues as well.
authors.affiliation: School of Pharmacy, Fudan University, Shanghai, PR China.
authors.forename: Hui
authors.initials: H
authors.lastname: Xie
authors.forename: Shengli
authors.initials: S
authors.lastname: Pan
authors.forename: Shufen
authors.initials: S
authors.lastname: Liu
authors.forename: Kan
authors.initials: K
authors.lastname: Ye
authors.forename: Keke
authors.initials: K
authors.lastname: Huo
mesh_terms: D029972:Bupleurum / Q000737:chemistry*; D002725:Chloroform / Q000737:chemistry; D015180:Electrophoresis, Gel, Two-Dimensional; D000439:Pentanols / Q000737:chemistry; D010936:Plant Extracts / Q000737:chemistry; D010940:Plant Proteins / Q000737:chemistry / Q000302:isolation & purification*; D018517:Plant Roots / Q000737:chemistry*; D014867:Water / Q000737:chemistry
chemical_list: C051274:Bupleurum root extract; D000439:Pentanols; D010936:Plant Extracts; D010940:Plant Proteins; D014867:Water; D002725:Chloroform; C029683:isopentyl alcohol