National Changhua University of Education Institutional Repository : Item 987654321/16767
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题名: Molecular Cloning of a cDNA Coding for Copper/Zinc Superoxide Dismutase from Zebrafish and Its Expression in Escherichia coli
作者: Ken, Chuian-Fu;Shaw, Jei-Fu;Wu, Jen-Leih;Lin, Chi-Tsai
贡献者: 生物學系
关键词: cDNA;Copper / zinc superoxide dismutase;Danio rerio;Escherichia coli;Expression;Molecular cloning;PCR;pET-23a(+)-thioredoxin;Zebrafish
日期: 1998-07
上传时间: 2013-06-05T08:48:06Z
出版者: American Chemical Society
摘要: A full-length complementary DNA (cDNA) clone encoding a putative copper/zinc Superoxide dismutase (Cu/Zn-SOD) was amplified by a polymerase chain reaction (PCR)-based technique from cDNA synthesized from zebrafish (Danio rerio) mRNA. Nucleotide sequence analysis of this cDNA clone revealed that it comprised a complete open reading frame coding for 154 amino acid residues. The deduced amino acid sequence showed higher identity (73.5-74.3%) with swordfish and shark Cu/Zn-SOD than with Cu/Zn-SOD from mammals (69.6-70.9%) and plants (55.8-56.2%). The ammo acid residues required for coordinating copper and zinc are conserved, as they are present in all reported Cu/Zn-SOD sequences. It lacks a targeting sequence, which suggests that the zebrafish cDNA clone encodes a cytosolic Cu/Zn-SOD. Furthermore, the coding region of Cu/Zn-SOD from zebrafish was introduced into an expression vector, pET-23a(+)-thioredoxin, and transformed into Escherichia coli AD494(DE3)pLysS. A predominant achromatic zone was detected by activity staining of native PAGE, and the expression pattern was shown by Coomassie blue staining of SDS-PAGE. This indicates that the Cu/Zn-SOD cDNA clone can be expressed in E. coli.
關聯: Journal of Agricultural and Food Chemistry, 46(7): 2863-2867
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