Recombinant inbred (RI) strains are derived from F2 individuals obtained by crossing of two highly inbred strains. Randomly chosen F2 pairs are brother-sister mated for more than 20 generations to become genetically fixed. Individual RI strains have unique combinations of loci derived by segregation and recombination of the alleles present in the progenitor strains. An important feature of RI strains is that, because they are inbred, repeated assays can be made so that phenotypes of each strain can be more precisely estimated using average results obtained from measurements in multiple rats per each RI strain. A precise estimation of phenotype-genotype relationship is especially important when studying highly variable traits such as blood pressure or behavior. Moreover, because data are cumulative across assays, studies, and research groups, accumulated data can be analyzed and trait relationships studied in a manner not possible with conventional, genetically segregating populations. This is a major advantage for the analysis of complex pathophysiologic traits.
The BXH/HXB RI strains were derived from reciprocal crosses of the SHR/Ola and BN-Lx
/Cub (Brown Norway) progenitors [28
]. At present, 20 HXB RI strains and 10 BXH RI strains are available (F>80). The RI strains were genotyped in over 13,000 SNPs that bin into 1,200 strain distribution patterns [29
]. In addition, over 130 copy number variants were determined in the RI strains [30
]. More than 200 physiological phenotypes determined in RI strains are available online in the GeneNetwork database for genetic analyses [31
]. The GeneNetwork database is a public resource that combines genomic and phenotypic information, including gene expression profiles, with fast software for linkage and correlation analyses [32
The BXH/HXB RI strains were used to map the genetic determinants of gene expression in key tissues relevant to pathophysiology of hypertension and the metabolic syndrome, including kidney, retroperitoneal fat, adrenals, left ventricles, whole brains, livers, soleus muscles, aorta, and hypocampus [23
] (Hubner, Williams, Pravenec, et al., unpublished results). After assessment of genome-wide significance and accounting for multiple testing using false discovery rates, over 1000 expression QTL (eQTL) were found in these tissues. Linkage analyses of gene expression profiles identified cis-
acting eQTL. These eQTL, especially cis-
regulated eQTL in the vicinity of physiological QTL (pQTL), represent a large source of attractive candidate genes for pQTL. In addition, gene expression profiles in relevant tissues can be correlated with physiological traits to reveal quantitative trait transcripts (QTT) and thus prioritize specific genes as candidates for discovery of specific QTGs. Using this strategy, Ogn
genes have been identified as QTGs predisposing to cardiac hypertrophy and hypertension, respectively [18
•]. Similarly, Ephx2
(epoxide hydrolase 2, cytoplasmic) has been identified as a genetic determinant predisposing SHHF (spontaneously hypertensive heart failure) rats to heart failure [36
]. Gene expression profiles thus represent useful intermediate phenotypes between variability at the DNA level and complex physiological traits. Additional intermediate phenotypes, including microRNA expression, quantitative proteome, metabonome, or epigenetic modifications (epigenome), will be determined in the BXH/HXB RI strains within the new integrated project EURATRANS, funded by the European Union [37
]. These new intermediate traits will provide new levels of ever-increasing complexity, which will enable investigators to build gene regulatory networks and associate these with complex physiological traits.