Is it possible to increase apospory and apomixis expressivity by hybridization and/or autopolyploidization in diploid Paspalum rufum?

DELGADO LUCIANA; ESPINOZA, FRANCISCO; MARIA E. SARTOR; MARIANO, SOLIMAN; GALDEANO, FLORENCIA; JUAN PABLO A. ORTIZ

Bahía Blanca

Workshop; V Ciclo de seminarios sobre avances en la caracterización genética y molecular de la apomixis; 2016

CERZOS-CONICET

Paspalum rufum Nees is a perennial grass that forms a multiploidcomplex composed of diploid (2n = 2x = 20) and tetraploid (2n = 4x = 40)cytotypes. The diploid form is sexual and highly self-sterile, while thetetraploid is a self-fertile pseudogamous aposporous apomict. Cytoembryologicalanalysis revealed that some diploid Paspalumcytotypes presented aposporous embryo sacs (AES) together with the meioticembryo sacs (MES) and also confirmed 5% of apomictic seeds produced in aself-pollination progeny. Subsequently, three natural populations ofdiploid P. rufum were analyzed,in order to evaluate the functionality of the apomixis component at the diploidlevel. This analysis allowed isolation of a single individual with capacity tocomplete apomixis (15% of the seed set), namely R5#49,  which was classified as ?diploid withresidual apomixis? and another one, R6#45, which produced the whole seed set bysexuality. Both individual were able to produce AES in 5.8 and 13 % of theirovules, respectively. The objective of this work were to characterize thereproductive behavior (evaluating both apospory and parthenogenesis) of diploidP. rufum hybrids from a segregant F1 population, and toanalyze the influence of hybridity and autopolyploidy on reproductive behavior.An F1 family of 39 individuals was created by crossing R6#45 x R5#49.Besides, two synthetic autotetraploids were obtained by colchicine treatment ofR6#45 mature seeds. The hybrid origin of the F1 progeny wasconfirmed by segregation analyses of a morphological trait and by the use of molecularmarkers. The ploidy level of experimental plants was determined by flowcytometry. The apospory expressivity was estimated by cytoembryologicalanalysis of cleared ovaries at anthesis. Out of the 39 hybrids analyzed, 38 formedAES. The expressivity of the trait ranged from 0 to 36 %, with some individualssignificantly exceeding the apospory expressivity values of the progenitors.Double-diploid plants had 25 and 32 % of its ovules carrying AES, respectively,a significantly higher rate comparing with that observed in R6#45. Then, we analyzedthe effect of hybridity and polyploidy on other apomixis components. Diploidhybrids producing different AES proportion were evaluated on their ability toproduce seeds through apomixis. Five individuals with apospory expressivitybetween 0 to 32% were induced to self-pollinate by using inter-specific,inter-ploidy pollen. The seeds DNA content was analyzed by flow cytometry,aimed at determining the embryo/endosperm DNA content ratio, which is typicalof the reproductive mode (2:3 for seed produced from sexuality; 2:5 for seedsproduced through apomixis). Experimental evidence indicated that most seedswere produced by selfing of MES. Only 2% and 0.8% of apomixis was detected intwo individuals with relatively high apospory expressivity. Meanwhile, the pollinationsystems of autopolyploids were evaluated; they still hold self-incompatibilityunder selfing pollination condition, producing a small seed set. Moreover, flowcytometry analysis revealed a very low capacity to produce seeds by apomixis.Only one seed of 169 analyzed was produced by apomixis. A second seed wasoriginated by cross pollination of an AES, generating a BIII hybrid. Our resultsrevealed a high variability in apospory expressivity of diploid hybrids, suggestingmulti-allelic and/or epigenetic control. The double-diploid reproductivecharacterization showed that apospory expressivity is highly hybridizationand/or ploidy dependent. Contrastingly, an increase in apospory expressivity associatedto hybridity or polyploidization is not sufficient to trigger parthenogenesis.