Identification and characterization of Clostridium perfringens using single target DNA microarray chip.
A DNA microarray method was developed to identify the presence of toxin genes: encoding beta toxin (cpb), epsilon toxin (etx), enterotoxin (cpe), alpha toxin (cpa), and iota toxin (iA) in Clostridium perfringens. To build the DNA chip, each gene sequence was represented by one approximately 22-bp amino-modified oligonucleotide printed twice on aldehyde-coated slides. Multiplex PCR with Cy3 and Cy5-dCTP derivatized fluorescent nucleotides was used to label five genes and fluorescent probes were prepared. The PCR probes were denatured and single-strand-labeled DNAs were separated and purified using magnetic beads. The presence of toxin genes in C. perfringens was detected by hybridization of amplified ssDNA probes to oligonucleotides on the chip representing one target sequence of each toxin gene. The DNA chip was able to identify eight strains of C. perfringens.
Al-Khaldi
Sufian F
SF
HFS-517, Division of Microbiological Studies, Center for Food Safety and Applied Nutrition, Food and Drug Administration, College Park, MD 20740-3855, USA. Sufian.Al-Khaldi@cfsan.fda.gov
Villanueva
Doralis
D
Chizhikov
Vladimir
V
D003016:Clostridium perfringens / Q000235:genetics / Q000302:isolation & purification*; D017931:DNA Primers; D004269:DNA, Bacterial / Q000737:chemistry*; D004768:Enterotoxins / Q000235:genetics*; D005506:Food Contamination; D005516:Food Microbiology; D005784:Gene Amplification; D005838:Genotype; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D016133:Polymerase Chain Reaction
D017931:DNA Primers; D004269:DNA, Bacterial; D004768:Enterotoxins