IL15多克隆抗体的制备及其在猪繁殖与呼吸综合征病毒活载体中的表达OA

This study aimed to prepare polyclonal antibodies against IL15 and validate their use with the plasmid pCDNA3.1-IL15 and the recombinant virus rPRRSV-IL15.Firstly,the plasmid pET-IL15 was constructed for prokaryotic expression with IPTG induction.The expressed recombinant protein was confirmed in SDS-PAGE to determine its reactivity and antigenicity.The purified recombinant IL15 protein was used to immunize rabbits to generate polyclonal antibodies.Meanwhile,the IL15 gene was inserted into the infectious clone of the highly pathogenic porcine reproductive and respiratory syndrome virus(PRRSV)strain vHuN4-F112 using reverse genetics techniques,resulting in the rescue of the recombinant virus rPRRSV-IL15,which stably expresses the IL15 protein.Subsequently,the plasmid pCDNA3.1-IL15 expressing IL15 protein was transfected into HEK 293T cells,and rPRRSV-IL15 infected MARC-145 cells and PAMs.The prepared antibodies were used to validate the IL15 expression in the transfected cells and rPRRSV-IL15-infected cells.The results demonstrated that the prepared polyclonal antibodies against IL15 showed good reactivity and specificity,capable of recognizing and binding to IL15 protein in the cells transfected with pCDNA3.1-IL15 and infected with rPRRSV-IL15.This study laid an important foundation for further investigating the biological functions of IL15 protein and its role in immune regulation.The recombinant virus rPRRSV-IL15 also represented an attempt to develop a novel immunoenhancing PRRSV attenuated vaccine.