title: RNA-Seq alignment to individualized genomes improves transcript abundance estimates in multiparent populations.
abstract: Massively parallel RNA sequencing (RNA-seq) has yielded a wealth of new insights into transcriptional regulation. A first step in the analysis of RNA-seq data is the alignment of short sequence reads to a common reference genome or transcriptome. Genetic variants that distinguish individual genomes from the reference sequence can cause reads to be misaligned, resulting in biased estimates of transcript abundance. Fine-tuning of read alignment algorithms does not correct this problem. We have developed Seqnature software to construct individualized diploid genomes and transcriptomes for multiparent populations and have implemented a complete analysis pipeline that incorporates other existing software tools. We demonstrate in simulated and real data sets that alignment to individualized transcriptomes increases read mapping accuracy, improves estimation of transcript abundance, and enables the direct estimation of allele-specific expression. Moreover, when applied to expression QTL mapping we find that our individualized alignment strategy corrects false-positive linkage signals and unmasks hidden associations. We recommend the use of individualized diploid genomes over reference sequence alignment for all applications of high-throughput sequencing technology in genetically diverse populations.
authors.affiliation: The Jackson Laboratory, Bar Harbor, Maine 04609.
authors.forename: Steven C
authors.identifier: http://orcid.org/0000-0002-8458-1871
authors.initials: SC
authors.lastname: Munger
authors.affiliation: The Jackson Laboratory, Bar Harbor, Maine 04609.
authors.forename: Narayanan
authors.initials: N
authors.lastname: Raghupathy
authors.affiliation: The Jackson Laboratory, Bar Harbor, Maine 04609.
authors.forename: Kwangbom
authors.initials: K
authors.lastname: Choi
authors.affiliation: The Jackson Laboratory, Bar Harbor, Maine 04609.
authors.forename: Allen K
authors.initials: AK
authors.lastname: Simons
authors.affiliation: The Jackson Laboratory, Bar Harbor, Maine 04609.
authors.forename: Daniel M
authors.initials: DM
authors.lastname: Gatti
authors.affiliation: The Jackson Laboratory, Bar Harbor, Maine 04609.
authors.forename: Douglas A
authors.initials: DA
authors.lastname: Hinerfeld
authors.affiliation: The Jackson Laboratory, Bar Harbor, Maine 04609.
authors.forename: Karen L
authors.initials: KL
authors.lastname: Svenson
authors.affiliation: University of Wisconsin, Madison, Wisconsin 53705.
authors.forename: Mark P
authors.initials: MP
authors.lastname: Keller
authors.affiliation: University of Wisconsin, Madison, Wisconsin 53705.
authors.forename: Alan D
authors.initials: AD
authors.lastname: Attie
authors.affiliation: The Jackson Laboratory, Bar Harbor, Maine 04609 Trinity University, San Antonio, Texas 78212.
authors.forename: Matthew A
authors.initials: MA
authors.lastname: Hibbs
authors.affiliation: The Jackson Laboratory, Bar Harbor, Maine 04609.
authors.forename: Joel H
authors.initials: JH
authors.lastname: Graber
authors.affiliation: The Jackson Laboratory, Bar Harbor, Maine 04609.
authors.forename: Elissa J
authors.initials: EJ
authors.lastname: Chesler
authors.affiliation: The Jackson Laboratory, Bar Harbor, Maine 04609 gary.churchill@jax.org.
authors.forename: Gary A
authors.initials: GA
authors.lastname: Churchill
mesh_terms: D000818:Animals; D005260:Female; D016678:Genome; D008297:Male; D051379:Mice; D040641:Quantitative Trait Loci; D016415:Sequence Alignment / Q000379:methods*; D017423:Sequence Analysis, RNA / Q000379:methods*; D012984:Software; D059467:Transcriptome
keywords: Diversity Outbred (DO); Diversity Outbred mice; MPP; Multiparent Advanced Generation Inter-Cross (MAGIC); Multiparental populations; QTL mapping; RNA-seq; expression QTL; haplotype reconstruction; high-density genotyping; mixed models