{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,5,14]],"date-time":"2025-05-14T00:10:04Z","timestamp":1747181404227,"version":"3.40.5"},"reference-count":44,"publisher":"Microbiology Society","issue":"12","funder":[{"name":"The Innovation Research Team for Basic and Clinical Studies on Chronic Liver Diseases of 2018 High-Level Health Teams of Zhuhai"},{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"crossref","award":["81971938"],"award-info":[{"award-number":["81971938"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"crossref","award":["81901557"],"award-info":[{"award-number":["81901557"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/501100012166","name":"National Basic Research Program of China","doi-asserted-by":"crossref","award":["2015CB554301"],"award-info":[{"award-number":["2015CB554301"]}],"id":[{"id":"10.13039\/501100012166","id-type":"DOI","asserted-by":"crossref"}]},{"name":"Entrepreneurial Talent Team Award of Guangdong Province","award":["2016ZT06S252"],"award-info":[{"award-number":["2016ZT06S252"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2021,12,24]]},"abstract":"<jats:p>Hepatitis C virus (HCV) genotype 3 is widely distributed, and genotype 3-infected patients achieve a lower cure rate in direct-acting antiviral (DAA) therapy and are associated with a higher risk of hepatic steatosis than patients with other genotypes. Thus, the study of the virology and pathogenesis of genotype 3 HCV is increasingly relevant. Here, we developed a full-length infectious clone and a subgenomic replicon for the genotype 3a isolate, CH3a. From an infected serum, we constructed a full-length CH3a clone, however, it was nonviable in Huh7.5.1 cells. Next, we systematically adapted several intergenotypic recombinants containing Core-NS2 and 5\u2032UTR-NS5A from CH3a, and other sequences from a replication-competent genotype 2\u2009a clone JFH1. Adaptive mutations were identified, of which several combinations facilitated the replication of CH3a-JFH1 recombinants; however, they failed to adapt to the full-length CH3a and the recombinants containing CH3a NS5B. Thus, we attempted to separately adapt CH3a NS5B-3\u2032UTR by constructing an intragenotypic recombinant using 5\u2032UTR-NS5A from an infectious genotype 3a clone, DBN3acc, from which L3004P\/M in NS5B and a deletion of 11 nucleotides (\u039411nt) downstream of the polyU\/UC tract of the 3\u2032UTR were identified and demonstrated to efficiently improve virus production. Finally, we combined functional 5\u2032UTR-NS5A and NS5B-3\u2032UTR sequences that carried the selected mutations to generate full-length CH3a with 26 or 27 substitutions (CH3acc), and both revealed efficient replication and virus spread in transfected and infected cells, releasing HCV of 10<jats:sup>4.2<\/jats:sup> f.f.u. ml<jats:sup>\u22121<\/jats:sup>. CH3acc was inhibited by DAAs targeting NS3\/4A, NS5A and NS5B in a dose-dependent manner. The selected mutations permitted the development of subgenomic replicon CH3a-SGRep, by which L3004P, L3004M and \u039411nt were proven, together with a single-cycle virus production assay, to facilitate virus assembly, release, and RNA replication. CH3acc clones and CH3a-SGRep replicon provide new tools for the study of HCV genotype 3.<\/jats:p>","DOI":"10.1099\/jgv.0.001704","type":"journal-article","created":{"date-parts":[[2021,12,24]],"date-time":"2021-12-24T10:15:28Z","timestamp":1640340928000},"source":"Crossref","is-referenced-by-count":1,"title":["Development of full-length cell-culture infectious clone and subgenomic replicon for a genotype 3a isolate of hepatitis C virus"],"prefix":"10.1099","volume":"102","author":[{"given":"Mingxiao","family":"Chen","sequence":"first","affiliation":[{"name":"Joint Program in Pathology, Department of Internal Medicine, Guangzhou Women and Children\u2019s Medical Center, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou 510623, PR China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yi","family":"Xu","sequence":"additional","affiliation":[{"name":"Joint Program in Pathology, Department of Internal Medicine, Guangzhou Women and Children\u2019s Medical 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