Isolation and characterization of serine protease gene(s) from Perkinsus marinus

Diseases of Aquatic Organisms

Kimberly S. Reece¹  Gwynne D. Brown¹ 

关键词: Perkinsus marinus;    Subtilisin;    Serine protease;   

DOI  :  10.3354/dao057117

学科分类：生物科学（综合）

来源: Inter-Research

【 摘 要 】

ABSTRACT: This study reports the first serine protease gene(s) isolated from Perkinsus marinus.Using universal primers, a 518 bp subtilisin-like serine protease gene fragment was amplified from P. marinus genomic DNA and used as a probe toscreen a λ-phage P. marinus genomic library; 2 different λ-phage clones hybridized to the digoxigenin(DIG)-labeled subtilisin-like gene fragment. Following subcloning and sequencing of thelarger DNA fragment, a 1254 bp open reading frame was identified and later confirmed, by 5' and 3' random amplification of cDNA ends (RACE) and northern blot analysis, to contain the entire coding-region sequence.Sequence analysis of the 3' RACE resultsfrom 2 isolate cultures, VA-2 (P-1) and LA 10-1, revealed multiple polymorphic sites within and among isolates. We identified 2 different types of cDNA clones with 95.53% nucleotide sequence similarity, suggesting the possibility of 2 closely relatedgenes within the P. marinus genome. Southern blot analysis of genomic DNA from 12 genetically distinct P. marinus isolate cultures revealed 2 different banding patterns among isolates.

【 授权许可】

Unknown