Self-complementarity in adeno-associated virus enhances transduction and gene expression in mouse cochlear tissues
| Autor: | Bradley J Walters, Charles Askew, Andrew M. Davidoff, Graham R. Casey, R. Jude Samulski, Mark A. Brimble, Chengwen Li |
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| Rok vydání: | 2020 |
| Předmět: |
Male
Molecular biology viruses Gene Expression medicine.disease_cause Biochemistry Green fluorescent protein Viral Packaging Transduction (genetics) Mice Transduction Genetic Animal Cells Gene expression Medicine and Health Sciences Transgenes Adeno-associated virus Organ of Corti Neurons Multidisciplinary Dependovirus Cell biology Cochlea Inner Ear Outer Hair Cells Medicine Female Anatomy Cellular Types Genetic Engineering Research Article Science Genetic Vectors Green Fluorescent Proteins Biology DNA construction Serogroup Green Fluorescent Protein Microbiology Protein Domains In vivo Cell Line Tumor Virology medicine otorhinolaryngologic diseases Animals Biology and Life Sciences Proteins Afferent Neurons Genetic Therapy Cell Biology In vitro Viral Replication Mice Inbred C57BL Research and analysis methods Luminescent Proteins Molecular biology techniques Animals Newborn Lipofectamine Cell culture Ears Cellular Neuroscience Plasmid Construction Head Neuroscience |
| Zdroj: | PLoS ONE PLoS ONE, Vol 15, Iss 11, p e0242599 (2020) |
| ISSN: | 1932-6203 |
| Popis: | Sensorineural hearing loss is one of the most common disabilities worldwide. Such prevalence necessitates effective tools for studying the molecular workings of cochlear cells. One prominent and effective vector for expressing genes of interest in research models is adeno-associated virus (AAV). However, AAV efficacy in transducing cochlear cells can vary for a number of reasons including serotype, species, and methodology, and oftentimes requires high multiplicity of infection which can damage the sensory cells. Reports in other systems suggest multiple approaches can be used to enhance AAV transduction including self-complementary vector design and pharmacological inhibition of degradation. Here we produced AAV to drive green fluorescent protein (GFP) expression in explanted neonatal mouse cochleae. Treatment with eeyarestatin I, tyrphostin 23, or lipofectamine 2000 did not result in increased transduction, however, self-complementary vector design resulted in significantly more GFP positive cells when compared to single-stranded controls. Similarly, self-complementary AAV2 vectors demonstrated enhanced transduction efficiency compared to single stranded AAV2 when injected via the posterior semicircular canal, in vivo. Self-complementary vectors for AAV1, 8, and 9 serotypes also demonstrated robust GFP transduction in cochlear cells in vivo, though these were not directly compared to single stranded vectors. These findings suggest that second-strand synthesis may be a rate limiting step in AAV transduction of cochlear tissues and that self-complementary AAV can be used to effectively target large numbers of cochlear cells in vitro and in vivo. |
| Databáze: | OpenAIRE |
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