Gene expression study in diplosporous and sexual Eragrostis curvula genotypes with differing ploidy levels using ESTs and differential display

GERARDO D CERVIGNI; NORMA PANIEGO; SILVINA PESSINO; MARINA DÍAZ; JUAN PABLO SELVA; DIEGO ZAPPACOSTA; GERMÁN SPANGENBERG; VIVIANA ECHENIQUE

Sapporo, Japón

Simposio; 5th International Symposium on the Molecular Breeding of Forage and Turf (MBFT2007); 2007

The molecular nature of gene expression during the initiation and progress of diplosporous apomixis is  still unknown. Moreover, the basis of the close correlation between diplospory and polyploidy is not clarified yet. A comparative expression analysis was performed based on ESTs (expressed sequence tags) sequencing and differential display in near isogenic lines of  E. curvula, a diplosporous apomictic forage grass. A tetraploid genotype (T, 4x apo), a sexual diploid derivative obtained from tissue culture (D, 2x sex) and an artificial sexual tetraploid obtained from the diploid seeds after colchicine treatment (C, 4x sex) were used. A collection of ESTs was generated from 3 cDNA libraries derived from panicles of the near-isogenic lines and one from 12 days-old plant leaves. A total of 12,295 high-quality ESTs were clustered and assembled, rendering 8,884 unigenes (1,490 contigs and 7,394 singletons). A total of 7,029 (79.11%) unigenes were functionally categorized by BLASTX analysis against sequences deposited in public databases, but only 37.80 % could be classified according to Gene Ontology and 50% using the cereals gene index (GI). From the total of unigenes derived from the inflorescence libraries, 112 (1.26 %) showed significant differential expression in individuals with different ploidy level and/or variable reproductive mode. Independent comparisons between plants with different reproductive mode (same ploidy) or different ploidy level (same reproductive mode) allowed the identification of genes modulated in response to diplosporous development or polyploidization, respectively. A group of genes were differentially expressed or silenced only in the 4x sex plant, presenting similar levels of expression in the 4x apo and the 2x sex genotypes. Differential display analysis showed that, in general, the 4x apo and 4x sex expression profiles were more related and different from the 2x sex one, but confirmed the existence of this particular group of genes, in both inflorescences and leaves. The possible biological significance of this group of genes is that an increase in ploidy level requires the activation/silencing of it, but for some reason this activation/silencing fails in the diplosporous plant. It is tempting to hypothesize that the expression repatterning for  a number of genes during the ‘genetic diploidization’ process abort in diplosporous plants (the same expression pattern of the sexual diploid plant is maintained), and maybe diplospory itself could be the consequence of this failure. The high ratio of non categorized genes keeps open the possibility of finding new genes involved in diplospory and its regulation. The 254 EST-SSRs and 190 SNPs detected in this study will provide valuable resources for mapping the region associated to diplospory in Eragrostis.