ArticleHuman gene therapy2013
High efficiency restriction enzyme-free linear amplification-mediated polymerase chain reaction approach for tracking lentiviral integration sites does not abrogate retrieval bias.
Article in Human gene therapy, 2013. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 19 papers.
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Who cites it
19 citing papers in PubMed, 23 citations in OpenAlex.
- Measuring Proviral HIV-1 DNA: Hurdles and Improvements to an Assay Monitoring Integration Events Utilising HumanLife (Basel, Switzerland) · 2021Article
- Clonal tracking using embedded viral barcoding and high-throughput sequencing.Nature protocols · 2020Article
- Bidirectional Retroviral Integration Site PCR Methodology and Quantitative Data Analysis Workflow.Journal of visualized experiments : JoVE · 2017Article
- Reproducibility of Illumina platform deep sequencing errors allows accurate determination of DNA barcodes in cells.BMC bioinformatics · 2016Article
- A novel gammaretroviral shuttle vector insertional mutagenesis screen identifies SHARPIN as a breast cancer metastasis gene and prognostic biomarker.Oncotarget · 2015Article
- Lentiviral gene therapy using cellular promoters cures type 1 Gaucher disease in mice.Molecular therapy : the journal of the American Society of Gene Therapy · 2015Article
- Quantitative shearing linear amplification polymerase chain reaction: an improved method for quantifying lentiviral vector insertion sites in transplanted hematopoietic cell systems.Human gene therapy methods · 2015Article
- High-throughput monitoring of integration site clonality in preclinical and clinical gene therapy studies.Molecular therapy. Methods & clinical development · 2015Article
- Clonal Dominance With Retroviral Vector Insertions Near the ANGPT1 and ANGPT2 Genes in a Human Xenotransplant Mouse Model.Molecular therapy. Nucleic acids · 2014Article
- Lentiviral and targeted cellular barcoding reveals ongoing clonal dynamics of cell lines in vitro and in vivo.Genome biology · 2014Article
- Path to the clinic: assessment of iPSC-based cell therapies in vivo in a nonhuman primate model.Cell reports · 2014Article
- Development and Evaluation of Quality Metrics for Bioinformatics Analysis of Viral Insertion Site Data Generated Using High Throughput Sequencing.Biomedicines · 2014Article
- Clonal tracking of rhesus macaque hematopoiesis highlights a distinct lineage origin for natural killer cells.Cell stem cell · 2014Article
- Multiplexing clonality: combining RGB marking and genetic barcoding.Nucleic acids research · 2014Article
- Impact of next-generation sequencing error on analysis of barcoded plasmid libraries of known complexity and sequence.Nucleic acids research · 2014Article
- High-titer foamy virus vector transduction and integration sites of human CD34(+) cell-derived SCID-repopulating cells.Molecular therapy. Methods & clinical development · 2014Article
- Development of an inducible caspase-9 safety switch for pluripotent stem cell-based therapies.Molecular therapy. Methods & clinical development · 2014Article
- Review
- Evaluating a ligation-mediated PCR and pyrosequencing method for the detection of clonal contribution in polyclonal retrovirally transduced samples.Human gene therapy methods · 2013Article
Corrections and comments
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Authors and funding
6 authors at 1 institution in 1 country.
Funding
Abstract
Retroviral vectors are an efficient and widely employed means of introducing an exogenous expression cassette into target cells. These vectors have been shown to integrate semi-randomly into the cellular genome, and can be associated with genotoxicity due to impact on expression of proximate genes. Therefore, efficient and accurate integration site analysis, while quantifying contributions of individual vector-containing clones, is desirable. Linear amplification-mediated polymerase chain reaction (LAM-PCR) is a widely used technique for identifying integrated proviral and host genomic DNA junctions. However, LAM-PCR is subject to selection bias inherent in the reliance of the assay on the presence of a restriction enzyme-cutting site adjacent to a retrievable integration site, and it is further limited by an inability to discriminate prior to sequencing between the flanking genomic DNA of interest and uninformative internal vector DNA. We report a modified restriction enzyme-free LAM-PCR (Re-free LAM-PCR) approach that is less time and labor intensive compared to conventional LAM-PCR, but in contrast to some other nonrestrictive methods, compares in efficiency and sensitivity, excludes retrieval of uninformative internal vector sequences, and allows retrieval of integration sites unbiased by the presence of nearby restriction sites. However, we report that Re-free LAM-PCR remains inaccurate for quantitation of the relative contributions of individual integration site-containing clones in a polyclonal setting, suggesting that bias in LAM-PCR retrieval of integration sites is not wholly explained by restriction enzyme-related factors.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.