Partitioning transcript variation in Drosophila: abundance, isoforms, and alleles.

Multilevel analysis of transcription is facilitated by a new array design that includes modules for assessment of differential expression, isoform usage, and allelic imbalance in Drosophila. The ∼2.5 million feature chip incorporates a large number of controls, and it contains 18,769 3' expression probe sets and 61,919 exon probe sets with probe sequences from Drosophila melanogaster and 60,118 SNP probe sets focused on Drosophila simulans. An experiment in D. simulans identified genes differentially expressed between males and females (34% in the 3' expression module; 32% in the exon module). These proportions are consistent with previous reports, and there was good agreement (κ = 0.63) between the modules. Alternative isoform usage between the sexes was identified for 164 genes. The SNP module was verified with resequencing data. Concordance between resequencing and the chip design was greater than 99%. The design also proved apt in separating alleles based upon hybridization intensity. Concordance between the highest hybridization signals and the expected alleles in the genotype was greater than 96%. Intriguingly, allelic imbalance was detected for 37% of 6579 probe sets examined that contained heterozygous SNP loci. The large number of probes and multiple probe sets per gene in the 3' expression and exon modules allows the array to be used in D. melanogaster and in closely related species. The SNP module can be used for allele specific expression and genotyping of D. simulans.