DOI: 10.5061/dryad.fs4cp
title: Data from: Advanced characterization of DNA molecules in rAAV vector preparations by single-stranded virus next-generation sequencing

description: Recent successful clinical trials with recombinant adeno-associated viral
 vectors (rAAVs) have led to a renewed interest in gene therapy. However,
 despite extensive developments to improve vector-manufacturing processes,
 undesirable DNA contaminants in rAAV preparations remain a major safety
 concern. Indeed, the presence of DNA fragments containing antibiotic
 resistance genes, wild-type AAV, and packaging cell genomes has been found
 in previous studies using quantitative polymerase chain reaction (qPCR)
 analyses. However, because qPCR only provides a partial view of the DNA
 molecules in rAAV preparations, we developed a method based on
 next-generation sequencing (NGS) to extensively characterize
 single-stranded DNA virus preparations (SSV-Seq). In order to validate
 SSV-Seq, we analyzed three rAAV vector preparations produced by transient
 transfection of mammalian cells. Our data were consistent with qPCR
 results and showed a quasi-random distribution of contaminants originating
 from the packaging cells genome. Finally, we found single-nucleotide
 variants (SNVs) along the vector genome but no evidence of large
 deletions. Altogether, SSV-Seq could provide a characterization of DNA
 contaminants and a map of the rAAV genome with unprecedented resolution
 and exhaustiveness. We expect SSV-Seq to pave the way for a new generation
 of quality controls, guiding process development toward rAAV preparations
 of higher potency and with improved safety profiles.
description: Sample
 CorespondanceTable indicating the
 correspondence between samples and IDsChromosomal_distributionRead count in human chromosomes used for Figure 3ReferencesFasta and Genbank files containing the sequences and annotations for the references non-available in public repositories (Ad5_in_293_genome, Backbone-AAV-CMV-GFP-hTK, Cassette_AAV-CMV-GFP-hTK, pDP8_Kana, SSV9K2-CMV-GFP-HygroTK-bGHpA)VariantsVCF files and alternative allele counting files along rAAV genome, used for Figure 4 b.CoverageBED files containing the base per base sequencing coverage along rAAV genome + Summary table with normalized data. Used for Figure 4 a.Circos_DataData and configuration file used to generate the Circos plot. Used for figure 5ContaVect_Configuration_FilesConfiguration files for each sample used to run ContaVectRUN1_S4_R1.fastqRaw fastq file containing data for sample id RUN1_S4 R1RUN1_S1_R1.fastq.gzRUN1_S4_R2.fastqRaw fastq file containing data for sample id RUN1_S4 R2RUN1_S1_R1.fastqRaw fastq file containing data for sample id RUN1_S1 R1RUN1_S1_R2.fastqRaw fastq file containing data for sample id RUN1_S1 R2RUN1_S2_R1.fastqRaw fastq file containing data for sample id RUN1_S2 R1RUN1_S2_R2.fastqRaw fastq file containing data for sample id RUN1_S2 R2RUN1_S3_R1.fastqRaw fastq file containing data for sample id RUN1_S3 R1RUN1_S3_R2.fastqRaw fastq file containing data for sample id RUN1_S3 R2RUN1_S5_R1.fastqRaw fastq file containing data for sample id RUN1_S5 R1RUN1_S5_R2.fastqRaw fastq file containing data for sample id RUN1_S5 R2RUN1_S6_R1.fastqRaw fastq file containing data for sample id RUN1_S6 R1RUN1_S6_R2.fastqRaw fastq file containing data for sample id RUN1_S6 R2RUN1_S7_R1.fastqRaw fastq file containing data for sample id RUN1_S7 R1RUN1_S7_R2.fastqRaw fastq file containing data for sample id RUN1_S7 R2RUN1_S8_R1.fastqRaw fastq file containing data for sample id RUN1_S8 R1RUN1_S8_R2.fastqRaw fastq file containing data for sample id RUN1_S8 R2RUN2_S1_R1.fastqRaw fastq file containing data for sample id RUN2_S1 R1RUN2_S1_R2.fastqRaw fastq file containing data for sample id RUN2_S1 R2RUN2_S2_R1.fastqRaw fastq file containing data for sample id RUN2_S2 R1RUN2_S2_R2.fastqRaw fastq file containing data for sample id RUN2_S2 R2RUN2_S3_R1.fastqRaw fastq file containing data for sample id RUN2_S3 R1RUN2_S3_R2.fastqRaw fastq file containing data for sample id RUN2_S3 R2RUN2_S4_R1.fastqRaw fastq file containing data for sample id RUN2_S4 R1RUN2_S4_R2.fastqRaw fastq file containing data for sample id RUN2_S4 R2RUN2_S5_R1.fastqRaw fastq file containing data for sample id RUN2_S5 R1RUN2_S5_R2.fastqRaw fastq file containing data for sample id RUN2_S5 R2RUN2_S6_R1.fastqRaw fastq file containing data for sample id RUN2_S6 R1RUN2_S6_R2.fastqRaw fastq file containing data for sample id RUN2_S6 R2RUN2_S7_R1.fastqRaw fastq file containing data for sample id RUN2_S7 R1RUN2_S7_R2.fastqRaw fastq file containing data for sample id RUN2_S7 R2RUN2_S8_R1.fastqRaw fastq file containing data for sample id RUN2_S8 R1RUN2_S8_R2.fastqRaw fastq file containing data for sample id RUN2_S8 R2