The Distribution Of Gene Chip For Three Kinds Of Avian Immunosuppressive Diseases

In recent years, Avian immunosuppressive diseases have spread rapidly in Chinese poultry andseriously harmed poultry husbandry. avian immunosuppressive diseases have no effective commercialvaccine or drugs to prevention and treatment. Pathogen detection is important to purification of flocks. Inthis study, a gene chip assay was developed to detect chicken infectious anemia virus, reticuloendotheliosisvirus and avian leukosis virus subgroups A, C and D based on the multiplex PCR.The reference sequences of CAV, REV, ALV subgroups A, C and D were obtained from GenBank,Three pairs of primers of those genes were designed and synthesized, the downstream primers were labeledby Cy3. Then, multiplex polymerase chain reaction system was built. In addition, Ten probes of thosegenes were designed and synthesized based on the conservative region of sequences. According tothe designing matrix, the probes were spotted to the glass substrate using the chip microarrayer. The totalDNA was extracted from several viruses. The products which amplified by multiplex PCR using thelabeled primers were hybridized with the gene chip. The hybridization signal was detected and analyzed bya fluorescence scanner.After hybridization, the chip can simultaneously detect the chicken infectious anemia virus andavian reticuloendotheliosis virus, and distinguish the avian leukosis virus subgroups A, C and D. Thedetection sensitivity can be reached to107copies/mL using specific test. The chip has higher specificity.Comparing of the450clinical samples results using ELISA and gene chip, the detection and accuracy rateof gene chip were higher. This study shows that the chip can simultaneously detects CAV and REV, anddistinguish between ALV subgroups A, C and D. This method will provides potential for rapid surveillanceand differential diagnosis of CAV, REV and ALV subgroups A, C and D.We constructed avian leukemia virus gene chip with this research method. The wholeblood, serum, egg white and cloacal swabs of the same chicken were detected by ALV gene chip. Theresults show that the positive percentage of whole blood samples was69.23%by gene chip. Thepositive percentage of ALV-p27antigen was23.08%in egg albumen samples by commercialELISA kit. The detection rate of the chip is better than ELISA kit, and the whole blood isthe optimal material for detection of ALV gene chip.