Herpes simplex virus type 1 (HSV-1) amplicon vectors are promising gene delivery tools, but their utility in gene therapy has been impeded to some extent by their inability to achieve stable transgene expression. In this study, we examined the possibility of improving transduction stability in cultured human cells via site-specific genomic integration mediated by adeno-associated virus (AAV) Rep and inverted terminal repeats (ITRs). A rep(-) HSV/AAV hybrid amplicon vector was made by inserting a transgene cassette flanked with AAV ITRs into an HSV-1 amplicon backbone, and a rep(+) HSV/AAV hybrid amplicon was made by inserting rep68/78 outside the rep(-) vector 3' AAV ITR sequence. Both vectors also had a pair of loxP sites flanking the ITRs. The resulting hybrid amplicon vectors were successfully packaged and compared to a standard amplicon vector for stable transduction frequency (STF) in human 293 and Gli36 cell lines and primary myoblasts. The rep(+), but not the rep(-), hybrid vector improved STF in all three types of cells; 84% of Gli36 and 40% of 293 stable clones transduced by the rep(+) hybrid vector integrated the transgene into the AAVS1 site. Due to the difficulty in expanding primary myoblasts, we did not assess site-specific integration in these cells. A strategy to attempt further improvement of STF by "deconcatenating" the hybrid amplicon DNA via Cre-loxP recombination was tested, but it did not increase STF. These data demonstrate that introducing the integrating elements of AAV into HSV-1 amplicon vectors can significantly improve their ability to achieve stable gene transduction by conferring the AAV-like capability of site-specific genomic integration in dividing cells.

Download full-text PDF

Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC136298PMC
http://dx.doi.org/10.1128/jvi.76.14.7150-7162.2002DOI Listing

Publication Analysis

Top Keywords

hybrid amplicon
20
amplicon vector
12
hsv-1 amplicon
12
amplicon vectors
12
amplicon
9
herpes simplex
8
simplex virus
8
virus type
8
rep+ hybrid
8
transgene expression
8

Similar Publications

The emergence of liquid biopsy technologies holds great promise in the cancer setting, including in pediatric central nervous system (CNS) tumors. In contrast to broad lower-depth sequencing, commonly referred to as low pass whole genome sequencing (WGS), targeted platforms with a higher depth of coverage have also been established. Here, we review targeted liquid biopsy techniques with applicability to pediatric CNS tumors.

View Article and Find Full Text PDF

Evaluation of a next generation sequencing assay for Hepatitis B antiviral drug resistance on the oxford nanopore system.

J Clin Virol

January 2025

Division of Medical Microbiology and Virology, St. Paul's Hospital, Providence Health Care, Vancouver, British Columbia, Canada; Department of Pathology and Laboratory Medicine, University of British Columbia, Vancouver, British Columbia, Canada. Electronic address:

Background: Next-generation sequencing (NGS) for Hepatitis B virus (HBV) antiviral resistance (AVR) testing is a highly sensitive diagnostic method, able to detect low-level mutant subpopulations. Our clinical virology laboratory previously transitioned from DNA hybridization (INNO-LiPA) to NGS, initially with the GS Junior System and subsequently the MiSeq. The Oxford Nanopore Technology (ONT) sequencing system was evaluated for HBV resistance testing, with regards to sequencing accuracy and turn-around time.

View Article and Find Full Text PDF

Powdery mildew, caused by the fungus , is one of the primary causes of grape yield loss across the globe. While numerous resistance loci have been identified in various grapevine species, the genetic determinants of susceptibility to remain largely unexplored. Understanding the genetics of susceptibility for pathogenesis is equally important for developing durable resistance grapevines against this pathogen.

View Article and Find Full Text PDF

Introduction: MET amplification (METamp) can be a de novo or acquired resistance driver; however, the definition of METamp that best captures patients who may respond to targeted therapy remains debated. We explored the genomic landscape of METamp NSCLC including degree of amplification, co-drivers, amplicon size, and outcomes to MET inhibitors.

Methods: Hybrid-capture NGS-based genomic profiling from 88,547 tissue and 12,428 liquid NSCLC samples were queried for METamp (copy number (CN) ≥ ploidy + 4, or amplification ratio (AmpRatio; [CN/sample ploidy] ≥ 3).

View Article and Find Full Text PDF

Marker-assisted selection in segregating populations of tomatoes for resistance to TYLCV, ToMV, and Fusarium wilt.

Mol Biol Rep

January 2025

Department of Agronomy and Plant Breeding Sciences, Agricultural College of Aburaihan, University of Tehran, Pakdasht, Iran.

Background: Tomato yellow leaf curl virus (TYLCV), tomato mosaic virus (ToMV), and Fusarium wilt are three of tomatoes' most important viral and fungal diseases.

Methods And Results: In this study, the application of molecular markers associated with tomato yellow leaf curl virus resistance gene (Ty1), tomato mosaic virus resistance gene (Tm2), and Fusarium wilt resistance gene (I-1) (linked marker) were evaluated. In order to optimize and use SNP markers (by HRM diagnostic method) and SCAR markers, segregating populations of tomatoes were produced by self-pollination of commercial hybrid cultivars.

View Article and Find Full Text PDF

Want AI Summaries of new PubMed Abstracts delivered to your In-box?

Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!