Isolation and Properties of Recombinant Human High Mobility Group I/Y Chromosomal Protein
作者:
期刊:
Biotechnology & Biotechnological Equipment
(Taylor Available online 1997)
卷期:
Volume 11,
issue 3-4
页码: 46-50
ISSN:1310-2818
年代: 1997
DOI:10.1080/13102818.1997.10818941
出版商: Taylor & Francis
数据来源: Taylor
摘要:
Recombinant human high mobility group chromosomal protein I/Y has been expressed in E. coli BL21(DE3) strain using the expression system of Studier et al. (1990). Cells were transformed with bacterial plasmid pET15bHMG-I/Y containing cDNA of the human HMG I/Y protein inserted in-frame with a sequence coding six cnosecutive histidines at the N-terminus of the protein (Thanos and Maniatis, 1992). The recombinant protein has been isolated from the total protein extract by selective binding of the 6xHis-tag on Ni2+-NTA resin (Qiagen) using metalo belate affinty chromatography. The protein was tested by SDS polyacrylamide gel electrophoresis and immunochemical detection with anti rat brain HMG I antibodies (IgG fraction). The immunoperoxidase test was performed on Western blots with 4-chloro-l-naphtol/H2O2as a substrate and on dot blots with luminol as an enhanced hemiluminiscence emitting substrate (ECL—Amersham Life Sci.). By its electrophoretic and immunochemical reactions the obtained recombinant human HMG I/Y protein was identical with a natural HCIO4extracted rat brain HMG I chromosomal protein.
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