  RNA sequencing (RNA-seq) enables characterization and quantification of
individual transcriptomes as well as detection of patterns of allelic
expression and alternative splicing. Current RNA-seq protocols depend on
high-throughput short-read sequencing of cDNA. However, as ongoing advances are
rapidly yielding increasing read lengths, a technical hurdle remains in
identifying the degree to which differences in read length influence various
transcriptome analyses. In this study, we generated two paired-end RNA-seq
datasets of differing read lengths (2x75 bp and 2x262 bp) for lymphoblastoid
cell line GM12878 and compared the effect of read length on transcriptome
analyses, including read-mapping performance, gene and transcript
quantification, and detection of allele-specific expression (ASE) and
allele-specific alternative splicing (ASAS) patterns. Our results indicate
that, while the current long-read protocol is considerably more expensive than
short-read sequencing, there are important benefits that can only be achieved
with longer read length, including lower mapping bias and reduced ambiguity in
assigning reads to genomic elements, such as mRNA transcript. We show that
these benefits ultimately lead to improved detection of cis-acting regulatory
and splicing variation effects within individuals.
