High-throughput sequencing for the analysis of genomic DNA and gene expression in Populus spp.

The genus Populus is an important crop and a model system to understand molecular processes of growth, development, and responses to environmental stimuli in trees. Moreover the entire genome of Populus trichocarpa was sequenced. The aim of this research was studying genomic variation and evolution in the poplar genus, and the effects of such variations in producing heterosis in two interspecific hybrids between Populus deltoides and P. nigra. Heterosis, intended as the superior performance of hybrid progeny compared to their inbred parents, has been one of the driving forces in poplar breeding. The two interspecific hybrids used in our experiments exhibit different levels of heterosis, i.e., their productivity is for one genotype much larger than that of parents and, for the other genotype, is similar to that of parents. The molecular bases of heterosis are still to be fully clarified, though it appears that variations in intergenic regions can have a role in the heterotic phenotype. Hence, we studied the extent of variation in the repetitive component of the genome (especially retrotransposons) and its possible consequences on gene and allelic expression. During this research, bioinformatic and genomic analyses were performed aiming i) to characterize the repetitive component of the poplar genome, by the isolation and characterization of LTR-retrotransposons in the P. trichocarpa genome, and the production of a database of such elements; moreover the previously undescribed structure of poplar centromeres was evaluated by means of NGS techniques; ii) to analyze poplar genome repetitive component and its expression, studying, by Illumina RNAseq, the transcription of previously isolated LTR-retrotransposons, in control and drought stressed plants; iii) to study the poplar transcriptome, also in relation to drought and, for an indirect evaluation of cis-regulatory sequence variation in the poplar hybrid, to the differential expression between alleles in genes expressed in control and drought stress. Concerning LTR-retrotransposons, we observed a relatively recent burst of retrotransposons activity, though counterbalanced by high levels of DNA loss. A huge fraction of retrotransposons belong to unknown superfamilies, i.e. they are non-autonomous retrotransposons because lacking coding capacity. These elements are especially expressed in poplars. We also individuated two distinct centromeric repeats, that occur in all three analysed poplar species. Gene expression was analysed mapping RNAseq data to the complete poplar transcriptome, and a reference expression dataset was established. In several instances, the two alleles in a hybrid are flanked by different DNA sequences, affecting tissue specificity or temporal regulation of expression of genes. We found allele specific expression in many of 200 randomly chosen genes in different stress conditions. This suggests a differential role for the two alleles during hybrid growth and in its interaction with the environment. It is possible that the functional diversity of the two parental alleles in the hybrid may have an impact on hybrid performance through allelic complementation