Novel lysine biosynthetic gene sequences (LYS1 and LYS5) used as PCR targets for the detection of the pathogenic Candida yeast.
We report here a sensitive and specific polymerase chain reaction (PCR) detection assay for the pathogenic Candida yeast based on the novel LYS1 [encoding saccharopine dehydrogenase (SDH)] and LYS5 [encoding phosphopantetheinyl transferase (PPTase)] gene sequences of the fungal unique lysine biosynthetic pathway. Both LYS1 and LYS5 DNA-specific PCR primers SG1, SG2 and SG3, SG4, respectively, amplified predicted 483 and 648-bp fragments from Candida albicans genomic DNA but not from other selected fungal, bacterial, or human DNA. The 18S rDNA control primers exhibited positive amplifications in all PCR assays. The LYS1-and LYS5-specific primers strongly amplified C. albicans and Candida tropicalis target sequences; however, the LYS1 primers also weakly amplified fragments from Candida kefyr and Candida lusitaniae DNA. Both sets of primers amplified target sequences from less than 10 pg of serially diluted C. albicans DNA, and the LYS1 specific primers also detected DNA isolated from serially diluted 50 C. albicans cells. The PCR primers reported here are sufficiently sensitive and specific for the potential early detection of Candida infections with no possibility of false positive results from cross-contamination with bacterial or human DNA.
Guo
Shujuan
S
Department of Microbiology, Pearson Hall, Room 32, Miami University, Oxford, OH 45056, USA.
Bhattacharjee
J K
JK
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D002175:Candida / Q000145:classification / Q000235:genetics* / Q000378:metabolism; D017931:DNA Primers / Q000235:genetics; D004587:Electrophoresis, Agar Gel; D008239:Lysine / Q000096:biosynthesis*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D012443:Saccharopine Dehydrogenases / Q000235:genetics* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D017855:Transferases (Other Substituted Phosphate Groups) / Q000235:genetics* / Q000378:metabolism
D001426:Bacterial Proteins; D017931:DNA Primers; C082662:phosphopantetheinyl transferase; D012443:Saccharopine Dehydrogenases; D017855:Transferases (Other Substituted Phosphate Groups); D008239:Lysine