Integrated long- and short-read single-cell RNA-sequencing

Single-cell RNA sequencing analysis has recently provided snapshots of gene expression of specific cell types and enabled cell types classification within an heterogenous population. As well as transcriptional changes, alternative splicing events and modifications of components of splicing machinery actively contributes in shaping cellular phenotype as well as ageing process and diseases occurence. Current high-throughput single-cell RNA sequencing methods may lack information on cell-specific isoform expression, missing key aspects of cell biology. In the present work we introduce a novel approach using the 10X Genomics Chromium to generate short-read (Illumina) and long-read (Pacific Biosciences Sequel II) RNA-sequencing libraries from the same single cells. This approach produced single cell parallel transcriptional and splicing profiling that demonstrates for the first time cell-type specific isoform expression and alterations at transcriptional levels associated with ageing in haematopoietic stem and progenitor cells We generated single cell full length cDNA libraries using 10x Genomics platform from young and aged murine bone marrow. cDNA library has been then used to produce short read Illumina library following 10x protocol instructions, and simultanosly as an input for long read SMRT bell library preparation (PacBio). That approach enables sequencing of the full-length of the transcript and detection of alternative splicing or recombination events in addition to quantify gene expression changes Mouse 1 ( M26_....._fastq file for Illumina raw data, one PB Sequel I raw file and one PacBio Sequenl II raw file) Mouse 2 ( M27_...._fastq file for Illumina raw data, one PB Sequel I raw file and one PacBio Sequenl II raw file) Mouse 3 (M28_...._fastq file for Illumina raw data, one PB Sequel I raw file and one PacBio Sequenl II raw file) Mouse 4 (M29_...._fastq file for Illumina raw data, one PB Sequel I raw file and one PacBio Sequenl II raw file) Mouse 5 (M 30_...._fastq file for Illumina raw data, one PB Sequel I raw file and one PacBio Sequenl II raw file) Mouse 6 (M31_...._fastq file for Illumina raw data, one PB Sequel I raw file and one PacBio Sequenl II raw file)