Oxford Nanopore long-read sequencing has transformed genomics and transcriptomics by enabling direct, high-resolution analysis of RNA and DNA molecules, including structural variants, repetitive expansions, and epigenetic modifications. In the NeuroRNA group, we have applied this technology for several applications, including the quality control (QC) of lentiviral vectors, a critical step in gene therapy development.
In collaboration with VIVEbiotech and supported by the MedTech Initiative, we developed a robust assay to assess the genome integrity of lentiviral vectors. Both cDNA and direct RNA (dRNA) sequencing were applied to purified viral RNA from concentrated preparations, allowing comparison of sequencing efficiency, coverage, and fidelity across constructs of varying complexity.
Our analysis highlights the strengths and limitations of cDNA versus dRNA sequencing for vector characterization. We also demonstrate that modifying construct orientation and structure reduces alternative splicing, improving vector reliability.
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