Cubillos et al. BMC Genomics 2012, 13:117 http://www.biomedcentral.com/1471-2164/13/117 Background Most traits vary quantitatively between individuals and are significantly influenced by genetic variation and its interaction with the environment. In general, transcript abundance of a gene can be considered as a quantitative trait since it differs between individuals with respect to genetic factors [1,2]. The utilisation of technologies such as microarray and, lately, next generation sequencing, allowed the simultaneous quantification of thousands of transcripts, shedding light into the genetics behind gene expression variation [3-5]. Comparison in Arabidopsis thaliana showed that at least 46% of the genes in the genome are differentially expressed between a pair of accessions [6]. Variation in gene expression levels is highly heritable and specific genomic regions can be mapped, underpinning transcript abundance variation [1,3] and underlying phenotypic diversity, causing for example differential disease susceptibility in humans [7] or variation in metabolite accumulation in plants [8]... Previous studies in experimental segregating populations have proved the polygenic nature of gene expression variation among lines and mapped thousands of expres- sion quantitative trait loci (eQTLs) in different model organisms, e.g.: Arabidopsis thaliana [3,9-12], Saccharo- myces cerevisiae [13,14], and Caenorhabditis elegans [15]. No less important is the interaction between geno- types and the environment [16]. For instance, a third of the budding yeast genes showed a significant strain x condition interaction effect, demonstrating the complex- ity of the control of transcript abundance. Transcript level differences can originate from cis- and/or trans-regulatory changes. Cis- acting regulations represent polymorphisms in physical linkage to the gene, affecting the transcription in an allele-specific manner. Consequently, cis-eQTLs are usually detected as local-eQTLs, being mapped in the vicinity of the gene; whereas trans-acting regulations act distantly in a non allele-specific way and can be located anywhere in the genome, most likely detected as distant-eQTL [1,11,17]. Although linkage or association mapping approaches allow the identification of local- and distant- eQTLs, cis and trans effects should be directly assessed by allele-specific expression assays [18]. In yeast, most expression differences mapped as major trans-acting loci, where few modifiers regulate the expression of hundreds of genes [1,14]. Contrasting with these results, experiments in C. elegans revealed an opposite pattern, with an over-representation of local-eQTLs compared to distant regulators [15]. Only few similar studies have addressed this subject in A. thaliana. Using microarray technology in recombinant inbred lines (RILs) from a cross between accessions Bayreuth (Bay) and Shahdara (Sha), more than 36,000 eQTLs were described using Page 2 of 12 whole rosettes [3]. In contrast, a parallel study using Landsberg (Ler) and Cape Verde Island (Cvi) accessions as RIL progenitors, detected 9 times less eQTLs than those mapped in the Bay x Sha population [10]. Discre- pancies between the two studies were also evident in the ratio between local- and distant-eQTLs, likely due to differences in statistical power and significance thresholds [11]. Different growth conditions, stages and the lack of similar analytical strategies in these studies impairs fine comparisons between crosses, stressing the need for a systematic approach to better understand the role of natural variation in expression traits among Ara- bidopsis accessions. The identification of thousands of local eQTLs allows testing for marks of concerted evolution in transcript abundance from genes with related function [19]. Sev- eral studies have addressed this issue and demonstrated the presence of signatures of selection associated with expression traits in a number of systems [5,20]. For example, several pathways were identified in a yeast hybrid between two Saccharomyces species exhibiting polygenic directional divergence in cis-regulatory var- iants [5]. Similarly, evidence for selection in expression traits involved in processes such as growth, locomotion and memory was detected in two subspecies of Mus musculus [20]. Thus far, evidence for a directional allelic effect between Arabidopsis accessions is scarce. Tran- scriptome analyses for 18 natural accessions determined an overrepresentation of differentially expressed genes within GO categories, involving response to the biotic environment, among others [6]. In order to identify genomic regions underlying tran- script variation in A. thaliana and signatures of selection between genes with related function, we performed eQTL mapping in two well-established recombinant populations. The analysis was essentially based on a connected-cross design between three accessions [21]: the reference Columbia (Col-0), Cape Verde Island (Cvi-0) and Burren (Bur-0). Expression level variation was monitored in parti- cular for more than 26,166 annotated nuclear genes in 314 RILs and eQTLs were mapped using a novel R/eqtl pack- age, accurately designed for such a study. Here, we show that the eQTL landscape is moderately conserved across populations and many of the eQTLs identified are cross- specific. Moreover, we show that accounting for con- founding sources in one of the crosses increases the sensi- tivity and the power to detect eQTLs, allowing the identification of a potentially coordinated transcriptional evolution in genes with related functions. Results eQTL analyses were performed in three displays: two on young rosette transcripts in Cvi-0 × Col-0 (hereafter