A universal random DNA amplification and labeling strategy for microarray to detect multiple pathogens of aquatic animals

Authors：Junli Feng; Yi Wang; Renyao Jin; Guijie Hao

Source：Journal of Virological Methods, 2020.

DOI：10.1016/j.jviromet.2019.113761

• DNA microarray
• Aquatic pathogen
• Phi29 DNA polymerase
• Detection

Abstract(#br)Diseases caused by bacteria, fungi, and viruses pose a great threat to aquaculture. As DNA microarrays can be used to detect multiple pathogens, here we reported an array with the potential to simultaneously detect 13 bacterial and 11 viral pathogens of aquatic animals. The array included 853 oligonucleotide probes (20–40 mer) complementary to various virus-specific sequences and four chromosomal loci (16S rRNA, gyrB , dnaJ, and recA ) of bacteria. Multiplex PCR, phi29 DNA polymerase, and a Klenow fragment-based method were evaluated for amplifying and labeling the nucleic acid of pathogens. While array hybridization signals were most intense using pathogen sequences amplified by multiplex PCR, the phi29 DNA polymerase method was more convenient and ideal since it did not require sequence-specific primers that could bias against detecting novel pathogens. The feasibility of the phi29 DNA polymerase-based microarray strategy was also demonstrated by detecting multiple unknown pathogens from four samples of diseased fish and shrimps.