Rapid diagnostic methods for classifying avian leukosis subgroups in the field were needed for routine, large-scale screening. As a first step in method development, we inserted the avian leukosis virus subgroup A （A...Rapid diagnostic methods for classifying avian leukosis subgroups in the field were needed for routine, large-scale screening. As a first step in method development, we inserted the avian leukosis virus subgroup A （ALV-A） env gene into plasmid pcDNA3.1/Zeo （＋） and used this construct to transfect DF-1 cells. Zeocin-resistant cells were obtained after 2 weeks of zeocin selection. Then, the cells were analyzed using PCR, immunofluorescence, and Western blot for expression of the envA-encoded envelope protein after 30 serial passages. The DF-1/A cell line was completely resistant to 104 TCIDso/0.1 mL （50% tissue culture infective dose）ALV-A and was partially resistant to 10~ TCIDs0/0.1 mL ALV-A viral particles. By comparing the DF-1/A and DF-1 cell lines, an ALV-A isolate was identified using a gag-specific ELISAfor capsid protein p27. Thus, we established a DF-1/A cell line that was resistant to ALV-A infection. This cell line will be useful as a diagnostic tool.展开更多
基金The work was founded by the National Key R&D Program of China(2016YFD0501606)the Public Industry Research Program,the Ministry of Agriculture of China(201203055)+2 种基金the Program of Science and Technology Development of Guangdong Province,China(2015A020209145)the China Meat-Type Chicken Research System(CARS-42-G09)the Modern Agriculture Talents Support Program,Ministry of Agriculture of China([2012] no.160)
文摘Rapid diagnostic methods for classifying avian leukosis subgroups in the field were needed for routine, large-scale screening. As a first step in method development, we inserted the avian leukosis virus subgroup A （ALV-A） env gene into plasmid pcDNA3.1/Zeo （＋） and used this construct to transfect DF-1 cells. Zeocin-resistant cells were obtained after 2 weeks of zeocin selection. Then, the cells were analyzed using PCR, immunofluorescence, and Western blot for expression of the envA-encoded envelope protein after 30 serial passages. The DF-1/A cell line was completely resistant to 104 TCIDso/0.1 mL （50% tissue culture infective dose）ALV-A and was partially resistant to 10~ TCIDs0/0.1 mL ALV-A viral particles. By comparing the DF-1/A and DF-1 cell lines, an ALV-A isolate was identified using a gag-specific ELISAfor capsid protein p27. Thus, we established a DF-1/A cell line that was resistant to ALV-A infection. This cell line will be useful as a diagnostic tool.
