Products & Services
Applications

Applications

Nanopore sequencing offers advantages in all areas of research. Our offering includes DNA sequencing, as well as RNA and gene expression analysis and future technology for analysing proteins.

Learn about applications
View all Applications
Resources
News Explore
Contact
Back

Full-coverage native RNA sequencing of HIV-1 viruses

Publication

Date: 17th November 2019 | Source: BioRxiv

Authors: Alejandro R. Gener, Jason T. Kimata.

Objective: To evaluate native RNA sequencing for sequencing HIV-1 viral genomes.

Methods: Fifteen HIV-1 strains were processed with Direct RNA Sequencing (SQK-RNA002) library kits and sequenced on MinION Mk1B devices with RevD flow cells (Oxford Nanopore Technologies (ONT), Oxford, UK). Raw reads were converted to FASTQ, aligned to reference sequences, and assembled into contigs. Multi-sequence alignments of the contigs were generated and used for cladistics analysis.

Results: We sequenced full-length HIV-1 from the transcriptional start site to 3' LTR (100% virion genome) in 3 out of 15 isolates (89.6, NLAD8, AD17), achieving majority coverage (defined as > 50%) in another 7 out of 15 isolates. Inspection of NLAD8 sequence alignments revealed splicing or deletion signatures. Despite the strong 3′ bias, read coverage was sufficient to evaluate single-nucleotide variants (SNVs), insertions and deletions in 9 isolates, and to assemble HIV-1 genomes directly from viral RNA, achieving a maximum of 94% assembly coverage for NLAD8. Phylogenetic relationships were maintained at the level of contigs, as well as individual reads.

Conclusions: ONT native RNA sequencing performed as expected, covering full-length HIV-1 RNA without PCR or cDNA sequencing. Native single-molecule RNA sequencing supported previous models of HIV-1 replication, and samples exhibited strain-specific transcriptional signals. We propose Context Dependency Variant Classification to describe variants occurring in information-dense regions of HIV. These data provide a rich resource for emerging RNA modification detection schemes. Future work will expand HIV-1 transcript profiling to infection models and clinical samples.

Read the full text

Recommended for you

Open a chat to talk to our sales team
FAQs

FAQs

Search