Transcriptome profiling of embryonic development rate in rainbow trout advance backcross introgression lines

Embryonic development rate is a key trait significantly associated and genetically linked with both growth rate and sexual maturity in rainbow trout. To identify candidate genes underlying embryonic development rate, whole genome expression microarray analyses were conducted using embryos from a fourth generation backcross family; each backcross generation involved the introgression of the fast-developing alleles for the major development rate QTL (from the Clearwater clonal line [CW]) into a slow-developing clonal line (Oregon State University [OSU]). Embryos were collected at 15, 19, and 28 days post-fertilization. Microsatellite markers (One112ADFG, OMM1009 and OmyFGT12TUF) linked to the major embryonic development rate QTL region were used to determine the QTL genotype. The sex marker OMY-Y1 was used to determine the genotypic sex of each embryo. cDNA from 48 individual embryos were used for microarray expression analysis. An ANOVA modeling approach was used to detect differential gene expression between embryos possessing fast-developing genotypes and slow-developing genotypes. A total of 182 features have been identified with significant differences between embryonic development rate genotypes. Quantitative PCR was conducted on ten representative genes using the rainbow trout homologous sequences and microarray results were validated. Microarrays were co-hybridized with one sample labeled with Cy3 and another with Cy5 (dual channels). Hybridizations on each slide involved direct co-hybridization within time points (15, 19 and 28 dpf) between individuals of the same sex, but with different QTL genotypes (CW vs. OSU). A total of 24 microarray slides were hybridized by cDNA from 48 individual embryos, 8 slides for each development time point. Dye swap design was involved throughout the whole experiment.