en Molecular Immunology & Vaccines Molecular Immunology & Vaccines مقاله کامل Full Length/Original Article <div style="text-align: justify;"><span style="font-family:Times New Roman;"><span style="font-size:11pt"><span style="line-height:normal"><b><span style="font-size:10.5pt"><span style="color:black">Background:</span></span></b><span style="font-size:10.5pt"><span style="color:black"> Human group A rotaviruses (RVs), with an 11-segmented double-stranded RNA genome, are a cause of viral gastroenteritis in infants and young children. Despite the availability of effective live-attenuated vaccines, RV infection is still a global health challenge. Hence, a reverse genetics system enabling the manipulation of RV segments, such as genome reassortment, is essential for developing new RV vaccines. Herein, a reverse genetics system was used to rescue a reassortant human-bovine RV, incorporating the VP7 gene segment from the human RV strain Wa (serotype G1) and a recombinant RV.</span></span></span></span><br> <span style="font-size:11pt"><span style="line-height:normal"><b><span style="font-size:10.5pt"><span style="color:black">Methods: </span></span></b><span style="font-size:10.5pt"><span style="color:black">To rescue a reassortant human-bovine RV, we used a reverse genetics system in which 13 plasmid constructs were co-transfected into BHK-T7 cells. In this method, 10 plasmids were derived from bovine RV, one encoded the human RV VP7 segment of serotype G1, and two expressed the capping enzyme subunits of vaccinia virus. We also created a recombinant RV featuring enhanced green fluorescent protein (EGFP) by replacing the bovine NSP3 plasmid with an NSP3-EGFP construct, where EGFP expression was regulated by an internal ribosomal entry site.</span></span></span></span><br> <span style="font-size:11pt"><span style="line-height:normal"><b><span style="font-size:10.5pt"><span style="color:black">Results: </span></span></b><span style="font-size:10.5pt"><span style="color:black">Our results showed successful rescue of the reassortant human-bovine RV and the recombinant strain, with the NSP3-EGFP construct facilitating EGFP translation across multiple RV passages. Replacing the <i>NSP3</i> gene segment of bovine RV with the release of a recombinant RV enabled EGFP expression in several RV passages.</span></span></span></span><br> <b><span style="font-size:10.5pt"><span style="line-height:115%"><span style="color:black">Conclusion: </span></span></span></b><span style="font-size:10.5pt"><span style="line-height:115%"><span style="color:black">Our study establishes the feasibility of using a reverse&nbsp;genetics system to generate a live-attenuated RV vaccine candidate, potentially enhancing future control efforts against RV infections. </span></span></span></span></div> Genotype, Rotavirus, Vaccines 254 261 http://ibj.pasteur.ac.ir/browse.php?a_code=A-10-6900-1&slc_lang=en&sid=1 Mona Aminipour mona.ap2019@gmail.com 0009-0008-8152-2463 No Department of Microbiology, Karaj Branch, Islamic Azad University, Karaj, Iran Mohsen Lotfi mohlotfi2003@yahoo.com 0000-0001-9902-7999 No Department of Quality Control, Razi Vaccine and Serum Research Institute, Agricultural Research, Education and Extension Organisation (AREEO), Karaj, Iran Naser Harzandi nasharzan@yahoo.com 0000-0003-2783-4462 No Department of Microbiology, Ka.C, Islamic Azad University, Karaj, Iran Ebrahim Babapour 0000-0003-1882-9178 No Department of Microbiology, Ka.C, Islamic Azad University, Karaj, Iran Seyed Dawood Mousavi Nasab d.mosavinasab.1@gmail.com 0000-0002-4831-3108 No WHO Collaborating Center for Reference and Research on Rabies, Pasteur Institute of Iran, Tehran, Iran Mostafa Ghaderi ghaderi_viro@yahoo.com 0000-0002-8249-1231 Yes Department of Human Viral Vaccine, Razi Vaccine and Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran