产植酸酶菌株的筛选及其植酸酶基因的克隆

Screening of Phytase-producing Strain and Cloning of Phytase Gene from the Selected Strain

摘要:

从土壤中筛选到产植酸酶活性较高的烟曲霉菌株WY-2，其植酸酶最适pH为5.5，最适温度为55 ℃。通过对烟曲霉WY-2植酸酶基因进行PCR扩增，获得了一个1.5 kb大小的特异性产物，将其克隆到载体pMD18-T中。测序结果分析表明，该基因片段含有植酸酶基因完整的阅读框架 (ORF)，基因全长1459 bp，其中包含一个61 bp的内含子，编码465个氨基酸，有7个潜在的糖基化位点，5′端有一编码26个氨基酸的信号肽序列。该基因与已报道的烟曲霉ATCC34625植酸酶基因有91%同源性，编码的氨基酸序列同源性为91%。

Abstract:

A strain with relative higher phytase-producing ability, Aspergillus fumigatus WY-2 was screened from soil. The optimal pH and temperature for activity of the phytase from A.fumigatus WY-2 were 5.5 and 55 ℃, respectively. The gene encoding the phytase was amplified from genomic DNA of the strain by PCR, and a 1.5 kb DNA fragment was obtained and then was cloned into vector pMD18-T. The sequencing analysis revealed that the DNA fragment contained a whole open reading frame (ORF) of phytase gene. The phytase gene was 1459 bp in length included with a 61 bp intron and encoded 465 amino acids. A signal peptide encoding 26 amino acids was found at 5′end of the gene. There were 7 potential glycosylation sites in the phytase. The present phytase showed 91% identity in nucleotide sequence and 91% identity in deduced amino acids sequence to the previously reported A.fumigatus ATCC34625 phytase.