Single-cell transcriptomics in the context of long-read nanopore sequencing

Abstract

Single cell transcriptomics were declared twice ‘Method of the year’ by Nature Methods journal, in 2013 and 2019. It marks the beginning of new avenue in fundamental and clinical research, as the classic biochemical approach is already seen as limited, as pooling up the contents of a large number of cells together depicts an averaged image that is missing the decision maker cells. Short-read sequencing has already revolutionized biology and medicine, providing an unsurpassed accuracy of the reads, and now third generation long-read nanopore sequencing completes and extends transcriptomics with the ability to discover all full-length isoforms at a single-cell level, providing a deeper insight in our understating of physiology and pathology of tissue functions. This review provides insight on how nanopore technological specifics and limitations have been explored so far in implementing the only direct nucleic acid sequencing platform in single cells sequencing. We review how single-cell sequencing was introduced, most widely used conventional and microfluidic approaches for single cell isolation, ways for further sequencing library generation tailored for Illumina, and how these were adopted for the nanopore technology.