Cytological and molecular characterization of three gametoclones of Citrus clementina

Abstract Background Three gametoclonal plants of Citrus clementina Hort. ex Tan., cv. Nules, designated ESP, FRA, and ITA (derived from three labs in Spain, France, and Italy, respectively), were selected for cytological and molecular characterization in order to elucidate genomic rearrangements provoked by haploidization. The study included comparisons of their ploidy, homozygosity, genome integrity, and gene dosage, using chromosome counting, flow cytometry, SSR marker genotyping, and array-Comparative Genomic Hybridization (arrayCGH). Results Chromosome counting and flow cytometry revealed that ESP and FRA were haploid, but ITA was tri-haploid. Homozygous patterns, represented by a single peak (allele), were observed among the three plants at almost all SSR loci distributed across the entire diploid donor genome. Those few loci with extra peaks visualized as output from automated sequencing runs, generally low or ambiguous, might result from amplicons of paralogous members at the locus, non-specific sites, or unexpected recombinant alleles. No new alleles were found, suggesting the genomes remained stable and intact during gametogenesis and regeneration. The integrity of the haploid genome also was supported by array-CGH studies, in which genomic profiles were comparable to the diploid control. Conclusions The presence of few gene hybridization abnormalities, corroborated by gene dosage measurements, were hypothetically due to the segregation of hemizygous alleles and minor genomic rearrangements occurring during the haploidization procedure. In conclusion, these plants that are valuable genetic and breeding materials contain completely homozygous and essentially intact genomes

Variaci\uf3n de la tasa de enraizamiento asociada al n\ufamero de subcultivo y di\ue1metro de microtallos de casta\uf1o Castanea sativa Mill.

This study had the objective of determining the subculture number and the shoot basal diameter that produces the best rooting rate ex vitro of chestnut, Castanea sativa Mill., microshoots obtained via in vitro culturing. The plant material corresponded to microcuttings obtained from embryo cultures with between 7 and 12 subcultures in the proliferative stage, for which Driver and Kuniyuki Walnut (DKW) medium was used with the macronutrients reduced to a half and supplemented with 1 mg L-1 6-benzylaminopurine (BAP) and 0.1 mg L-1 indole-3-butiric acid (IBA). The quick induction rooting method was used, submerging the base of the microcuttings in a solution of 0.5 mg mL-1 of IBA for 15 min. For the rooting stage, pine bark:perlite (4:1, v/v) substrate was used, carrying out the evaluation of results at 20 days after culturing. The evaluated variables were survival rate (%), rooting rate (%), root number, root length (mm), callus presence and visual aspect of the rooting system. The results showed a decrease of rooting capacity as the subculture number increases. The factor microcutting diameter did not present significant differences with respect to the evaluated variables.Este estudio tuvo por objeto determinar el n\ufamero de subcultivo y el di\ue1metro basal en el cual se obtiene la mayor tasa de enraizamiento ex vitro para microtallos de casta\uf1o, Castanea sativa Mill., obtenidos v\ueda cultivo in vitro. El material vegetal correspondi\uf3 a microtallos provenientes del cultivo de embriones con entre 7 y 12 subcultivos durante la etapa de proliferaci\uf3n, para la cual se utiliz\uf3 medio Driver y Kuniyuki Walnut (DKW) con los macronutrientes reducidos a la mitad y suplementado con 1 mg L-1 6-benzilaminopurina (BAP) y 0,1 mg L-1 de \ue1cido indol 3-but\uedrico (AIB). Se utiliz\uf3 el m\ue9todo de inducci\uf3n r\ue1pida de enraizamiento sumergiendo los tallos en una soluci\uf3n de 0,5 mg mL-1 de AIB durante 15 min. Para la etapa de enraizamiento, se utiliz\uf3 como sustrato corteza de pino:perlita (4:1, v/v) evaluando los resultados a los 20 d\uedas de cultivo. Las variables evaluadas fueron tasa de supervivencia (%), tasa de enraizamiento (%), n\ufamero de ra\uedces, largo de ra\uedces (mm), presencia de callo y aspecto del sistema radicular. Los resultados mostraron una disminuci\uf3n de la capacidad de enraizamiento a medida que aumenta el n\ufamero de subcultivo. El factor di\ue1metro de microtallo no present\uf3 diferencias significativas respecto a las variables evaluadas

Dual regulation of water retention and cell growth by a stress-associated protein (SAP) gene in Prunus

We have identified a gene (PpSAP1) of Prunus persica coding for a stress-associated protein (SAP) containing Zn-finger domains A20 and AN1. SAPs have been described as regulators of the abiotic stress response in plant species, emerging as potential candidates for improvement of stress tolerance in plants. PpSAP1 was highly expressed in leaves and dormant buds, being down-regulated before bud dormancy release. PpSAP1 expression was moderately induced by water stresses and heat in buds. In addition, it was found that PpSAP1 strongly interacts with polyubiquitin proteins in the yeast two-hybrid system. The overexpression of PpSAP1 in transgenic plum plants led to alterations in leaf shape and an increase of water retention under drought stress. Moreover, we established that leaf morphological alterations were concomitant with a reduced cell size and down-regulation of genes involved in cell growth, such as GROWTH-REGULATING FACTOR (GRF)1-like, TONOPLAST INTRINSIC PROTEIN (TIP)-like, and TARGET OF RAPAMYCIN (TOR)-like. Especially, the inverse expression pattern of PpSAP1 and TOR-like in transgenic plum and peach buds suggests a role of PpSAP1 in cell expansion through the regulation of TOR pathway

Multipotent MAO and cholinesterase inhibitors for the treatment of Alzheimer's disease: synthesis, pharmacological analysis and molecular modeling of heterocyclic substituted alkyl and cycloalkyl propargyl amine.

The synthesis, pharmacological evaluation and molecular modeling of heterocyclic substituted alkyl and cycloalkyl propargyl amines 1-7 of type I, and 9-12 of type II, designed as multipotent inhibitors able to simultaneously inhibit monoamine oxidases (MAO-A/B) as well as cholinesterase (AChE/BuChE) enzymes, as potential drugs for the treatment of Alzheimer's disease, are described. Indole derivatives 1-7 of type I are well known MAO inhibitors whose capacity to inhibit AChE and BuChE was here investigated for the first time. As a result, compound 7 was identified as a MAO-B inhibitor (IC50 = 31 +/- 2 nM) and a moderately selective eqBuChE inhibitor (IC50 = 4.7 +/- 0.2 mu M). Conversely, the new and readily available 5-amino-7-(prop-2-yn-1-yl)-6,7,8,9-tetrahydropyrido[2,3-b][1,6]naphthyridine derivatives 9-13 of type II are poor MAO inhibitors, but showed AChE selective inhibition, compound 12 being the most attractive as it acts as a non-competitive inhibitor on EeAChE (IC50 = 25 +/- 3 nM, K-i = 65 nM). The ability of this compound to interact with the AChE peripheral binding site was confirmed by kinetic studies and by molecular modeling investigation. Studies on human ChEs confirmed that 12 is a selective AChE inhibitor with inhibitory potency in the submicromolar range. Moreover, in agreement with its mode of action, 12 was shown to be able to inhibit A beta aggregation induced by hAChE by 30.6%