A DE NOVO DIPLOID GENOME ASSEMBLY ALLOWS DISSECTING THE TRANSCRIPTOMIC DIFFERENCES UNDERLYING THE CLONAL PHENOTYPIC DIVERSITY IN CULTIVAR ‘MALBEC’

CALDERÓN LUCIANO; CARBONELL-BEJERANO, PABLO; MUÑOZ, CLAUDIO; BREE, LAURA; SOLA, CRISTOBAL; BERGAMIN, DANIEL; GOMEZ-TALQUENCA, SEBASTIAN; IBÁÑEZ, JAVIER; JOSÉ MIGUEL MARTÍNEZ-ZAPATER; LIJAVETZKY, DIEGO

Mendoza

Congreso; LVIII Annual Meeting of the Argentine Society for Biochemistry and Molecular Biology Research; 2022

SAIB

Most grapevine cultivars employed in the wine industry were originated from outcrossing two genetically divergent cultivars and then clonally propagated. Therefore, grapevine cultivars characterize by possessing diploid and highly heterozygous genomes. Despite the vegetative propagation system, intra-cultivar phenotypic and genetic diversity has been widely reported. Malbec is appreciated for producing high-quality red wines and recognized as the signature cultivar of the Argentine viticulture. Previous analyses of our group demonstrated notorious phenotypic differences among Malbec clones. Also, significant genetic differences were reported, mostly based on SNPs occurring across the intergenic regions. Here, aiming to dissect the transcriptomic bases of the clonal phenotypic diversity, we firstly assembled a truly-phased reference genome for Malbec. To perform the de novo assembly, we used a trio-binning approach implemented by the software Canu. We generated Illumina short-reads for Malbec parental cultivars (Magdeleine and Prunelard), that were employed to sort the PacBio long-reads generated for Malbec into its two component haplotypes. Finally, we obtained two assemblies: “Malbec-Prunelard” and “Malbec-Magdeleine”, each one showing the total length and gene content expected for a haploid grapevine genome. On the other hand, we studied 27 clonal accessions grown under the same conditions at Vivero Mercier experimental vineyard. Analytical and biochemical measurements were performed on mature berries (23º Brix), during two consecutive seasons (2017-2019). Afterwards, we chose the six clones exhibiting the greatest differences for the evaluated features. Whole RNA extractions were performed from berries to conduct RNA-seq experiments. More precisely, Illumina stranded paired-end reads were obtained for the six clones with their three biological replicates (18 samples). RNA-seq data was aligned to each haplotype of Malbec reference genome, to perform differential gene expression (DEG) and gene ontology (GO) enrichment analyses. After performing a multivariate discriminant analysis including all samples, clone 595 exhibited the most significant phenotypic and transcriptomic differences. In particular, 595 showed the highest total polyphenols concentration. At the same time, the DEG and GO enrichment analyses consistently showed that 595 exhibited significantly up-regulated genes involved in the anthocyanins biosynthesis pathways, when compared to the other clones. Furthermore, using a diploid assembly as reference enabled us to detect different sets of DEG and GO terms with each haplotype, and to observe differences in the intensity of the detected processes. Overall, these results highlight the importance of using a truly-phased assembly to analyze RNA-seq data and suggest that the clonal phenotypic diversity observed in Malbec could be explained by differences in the transcriptome.