title: Effect of read-mapping biases on detecting allele-specific expression from RNA-sequencing data.
abstract: BACKGROUND
  Next-generation sequencing has become an important tool for genome-wide quantification of DNA and RNA. However, a major technical hurdle lies in the need to map short sequence reads back to their correct locations in a reference genome. Here, we investigate the impact of SNP variation on the reliability of read-mapping in the context of detecting allele-specific expression (ASE).
  
  
  RESULTS
  We generated 16 million 35 bp reads from mRNA of each of two HapMap Yoruba individuals. When we mapped these reads to the human genome we found that, at heterozygous SNPs, there was a significant bias toward higher mapping rates of the allele in the reference sequence, compared with the alternative allele. Masking known SNP positions in the genome sequence eliminated the reference bias but, surprisingly, did not lead to more reliable results overall. We find that even after masking, approximately 5-10% of SNPs still have an inherent bias toward more effective mapping of one allele. Filtering out inherently biased SNPs removes 40% of the top signals of ASE. The remaining SNPs showing ASE are enriched in genes previously known to harbor cis-regulatory variation or known to show uniparental imprinting. Our results have implications for a variety of applications involving detection of alternate alleles from short-read sequence data.
  
  
  BACKGROUND
  Scripts, written in Perl and R, for simulating short reads, masking SNP variation in a reference genome and analyzing the simulation output are available upon request from JFD. Raw short read data were deposited in GEO (http://www.ncbi.nlm.nih.gov/geo/) under accession number GSE18156.
  
  
  BACKGROUND
  jdegner@uchicago.edu; marioni@uchicago.edu; gilad@uchicago.edu; pritch@uchicago.edu
  
  
  BACKGROUND
  Supplementary data are available at Bioinformatics online.
authors.affiliation: Department of Human Genetics, University of Chicago, 920 E. 58th St., CLSC 507, Chicago, IL 60637, USA. jdegner@uchicago.edu
authors.forename: Jacob F
authors.initials: JF
authors.lastname: Degner
authors.forename: John C
authors.initials: JC
authors.lastname: Marioni
authors.forename: Athma A
authors.initials: AA
authors.lastname: Pai
authors.forename: Joseph K
authors.initials: JK
authors.lastname: Pickrell
authors.forename: Everlyne
authors.initials: E
authors.lastname: Nkadori
authors.forename: Yoav
authors.initials: Y
authors.lastname: Gilad
authors.forename: Jonathan K
authors.initials: JK
authors.lastname: Pritchard
mesh_terms: D000483:Alleles; D001483:Base Sequence; D019295:Computational Biology / Q000379:methods*; D020869:Gene Expression Profiling; D015894:Genome, Human; D006801:Humans; D020641:Polymorphism, Single Nucleotide; D017423:Sequence Analysis, RNA / Q000379:methods*; D012984:Software