Novel recombinant papillomavirus genomes expressing selectable genes
| Autor: | Alison A. McBride, Wesley H. Stepp, Samuel S. Porter, Caleb McKinney, Koenraad Van Doorslaer |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2016 |
| Předmět: |
Gene Expression Regulation
Viral Genetic Markers Male 0301 basic medicine Genes Viral Virus Integration viruses Biology Early viral transcription Virus Replication Genome Article law.invention Colony-Forming Units Assay 03 medical and health sciences Viral life cycle law Extrachromosomal DNA Drug Resistance Viral Humans Gene Recombination Genetic Genetics Multidisciplinary Human papillomavirus 18 Infant Newborn Neomycin 030104 developmental biology Genetic marker Cell culture Recombinant DNA |
| Zdroj: | Scientific Reports |
| ISSN: | 2045-2322 |
| DOI: | 10.1038/srep37782 |
| Popis: | Papillomaviruses infect and replicate in keratinocytes, but viral proteins are initially expressed at low levels and there is no effective and quantitative method to determine the efficiency of infection on a cell-to-cell basis. Here we describe human papillomavirus (HPV) genomes that express marker proteins (antibiotic resistance genes and Green Fluorescent Protein), and can be used to elucidate early stages in HPV infection of primary keratinocytes. To generate these recombinant genomes, the late region of the oncogenic HPV18 genome was replaced by CpG free marker genes. Insertion of these exogenous genes did not affect early replication, and had only minimal effects on early viral transcription. When introduced into primary keratinocytes, the recombinant marker genomes gave rise to drug-resistant keratinocyte colonies and cell lines, which maintained the extrachromosomal recombinant genome long-term. Furthermore, the HPV18 “marker” genomes could be packaged into viral particles (quasivirions) and used to infect primary human keratinocytes in culture. This resulted in the outgrowth of drug-resistant keratinocyte colonies containing replicating HPV18 genomes. In summary, we describe HPV18 marker genomes that can be used to quantitatively investigate many aspects of the viral life cycle. |
| Databáze: | OpenAIRE |
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