Isolation and characterization of serine protease gene(s) from Perkinsus marinus.
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 to screen a lambda-phage P. marinus genomic library; 2 different lambda-phage clones hybridized to the digoxigenin(DIG)-labeled subtilisin-like gene fragment. Following subcloning and sequencing of the larger 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 results from 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 related genes 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.
Brown
Gwynne D
GD
Virginia Institute of Marine Science, PO Box 1346, Gloucester Point, Virginia 23062, USA.
Reece
Kimberly S
KS
D000595:Amino Acid Sequence; D000818:Animals; D010582:Bacteriophage lambda; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D003001:Cloning, Molecular; D016000:Cluster Analysis; D017931:DNA Primers; D056890:Eukaryota / Q000235:genetics*; D014644:Genetic Variation; D008969:Molecular Sequence Data; D010802:Phylogeny; D017422:Sequence Analysis, DNA; D012697:Serine Endopeptidases / Q000235:genetics*
D017931:DNA Primers; D012697:Serine Endopeptidases