麴菌銅鋅型超氧歧化酶基因選殖

Cloning and Characterization of a Copper/ Zinic-Superoxide Dismutase Gene from Aspergillus japonicus.

依據玉米Cu/Zn-SOD之核酸序列，合成兩股引子，並以麴菌cDNA當模板，以PCR方法增生出一段0.3 kb DNA，經次選殖及定序，其序列與玉米Cu/Zn-SOD之序列同殖性高達73%，進一步以此0.3 kb DNA當探針，進行麴菌基因庫之篩選，結果篩得一株含有0.4 kb之正反應株，經定序分析，發現可推導出含120個胺基酸序列，其序列和其它生物Cu/Zn-SOD之序列同質性很高，並含有與銅和鋅進行相對應（coordinating）之胺基酸序列。

In order to clone a Cu/Zn-SOD gene from Aspergillus japonicus, the highly conserved regions of the SOD sequence of maize and other species were compared and two oligonucleotides were synthesized. The Aspergillus japonicus cDNA was then used as the template to get a 0.3 kb fragment by PCR technique. The 0.3 kb DNA fragment has high homology compared with maize SOD-4 cDNA. Then we used the 0.3 kb fragment as probe to screen Aspergillus japonicus genomic libraries. A 0.4 kb DNA fragment from purified positive plaque was subcloned into pGEM-7zf(+) vector and sequenced. Nucleotide sequence analysis of this clone revealed that it comprised an open reading frame coding for 120 amino acid residues. The encoded polypeptide is highly homologous with other plant SODs. The residues required for coordinating copper and zinc are also conserved as they are among all reported Cu/Zn-SOD sequences.