| Popis: |
High-throughput short-read sequencing has taken on a central role in research and diagnostics. Literally hundreds of different assays exist today to take advantage of Illumina short-read sequencers, the predominant short-read sequencing technology available today. Although other short read sequencing technologies exist, the ubiquity of Illumina sequencers in sequencing core facilities, and the inertia associated with the research enterprise as a whole have limited their adoption. Among a new generation of sequencing technologies, Oxford Nanopore Technologies (ONT) holds a unique position because the ONT MinION, an error-prone long-read sequencer, is associated with little to no capital cost. Here we show that we can make short-read Illumina libraries compatible with the long-read ONT MinION by circularizing and rolling circle amplifying the short library molecules using the R2C2 method. This results in longer DNA molecules containing tandem repeats of the original short library molecules. This longer DNA is ideally suited for the ONT MinION, and after sequencing, the tandem repeats in the resulting raw reads can be converted into millions of high-accuracy consensus reads with similar error rates to that of the Illumina MiSeq. We highlight this capability by producing and benchmarking RNA-seq, ChIP-seq, as well as regular and target-enriched Tn5 libraries. We also explore the use of this approach for rapid evaluation of sequencing library metrics by implementing a real-time analysis workflow. |
