We extracted viral RNA from frozen serum samples from 11 untreated HIV patients, with approximately 8 time points per patient, spanning at least five years from an established time of infection. To maximize sensitivity, depth and coverage, we developed a protocol for whole-genome sequencing using six overlapping primer sets. We optimized the library preparation protocol to obtain long inserts starting from less than 1 ng of DNA and sequenced all samples on the Illumina MiSeq platform, obtaining around 10,000x coverage. We are able to call minor alleles as rare as 0.2% in the viral population and preserve linkage information over 500 bp.
This deep whole genome data set covering many years in 11 patients provides a comprehensive portrait of HIV-1 intra-patient evolution that should be suitable for many different types of analyses. We are planning to release the dataset soon and will in addition to raw reads provide pre-analyzed data such as consensus sequences, minor single nucleotide variant frequencies and tools to extract haplotypes up to 500 bp in length in most regions of the HIV-1 genome.
Population genomics of intrapatient HIV-1 evolution
Fabio Zanini, Johanna Brodin, Lina Thebo, Christa Lanz, Jan Albert and Richard Neher. eLIFE, e11282.