Selection of reference genes for gene expression studies in zucchini (Cucurbita pepo) using qPCR

The zucchini (Cucurbita pepo) is an important food crop, the transcriptomics of which are a fundamental tool to accelerate the development of new varieties by breeders. However, the suitability of reference genes for data normalization in zucchini has not yet been studied. The aim of this study was to assess the suitability of 13 genes for their potential use as reference genes in quantitative real-time PCR. Assays were performed on 34 cDNA samples representing plants under different stresses and at different developmental stages. The application of geNorm and NormFinder software revealed that the use of a combination of UFP, EF-1A, RPL36aA, PP2A, and CAC genes for the different experimental sets was the best strategy for reliable normalization. In contrast, 18S rRNA and TUA were less stable and unsuitable for use as internal controls. These results provide the possibility to allow more accurate use of qPCR in this horticultural crop. © 2011 American Chemical Society.Financial support was provided by the Spanish Project INIA-RTA2008-00035-C02-01, FEDER and FSE funds (EU).Peer Reviewe

Identification, cloning, and expression analysis of three phytoene synthase genes from Cucurbita pepo

An essential step in the carotenoid biosynthesis pathway is the formation of phytoene by phytoene synthase (PSY). In this study, three new genes coding PSYs (CpPSYA, CpPSYB, and CpPSYC) were cloned from Cucurbita pepo and their expression patterns analysed in three cultivars of summer squash which had a different carotenoid content. The gene sequences had a high similarity with those from other plant species, and their predicted proteins were significantly different from each other. A phylogenetic analysis indicates that CpPSYA and CpPSYB shared a high homology and were also homologous with PSYs from others cucurbits, whereas CpPSYC was more closely related to orthologues from strawberry and carrot. An expression analysis revealed that CpPSYA had a higher expression in flowers compared to leaves and showed a differential expression during fruit development. The amount of CpPSYA transcript was higher in fruits with a higher carotenoid content than in those with a lower carotenoid content. However, CpPSYB and CpPSYC showed a relatively high expression in leaves, and their expression in fruits varied among the different cultivars and fruit tissues. These results suggest that the CpPSY genes were under different regulatory mechanisms and they may have different roles in C. pepo.A financial support was provided by the Spanish Project INIA-RTA2011-00044-C02-01, FEDER and FSE funds. Á. Obrero had a contract from INIA-IFAPA (Subprograma FPI-INIA), co-financed by FSE funds (Programa Operativo FSE de Andalucía 2007–2013 “Andalucía se mueve con Europa”). The New Zealand Institute for Plant and Food Research Limited hosted Á. Obrero for a short stay, where part of this work was done. C.I. González Verdejo wish to thank the National Institute of Agricultural Research (INIA) and FSE funds for contract funding.Peer reviewe

Carotenogenic gene expression and carotenoid accumulation in three varieties of Cucurbita pepo during fruit development

The control of gene expression is a crucial regulatory mechanism in carotenoid accumulation of fruits and flowers. We investigated the role of transcriptional regulation of nine genes involved in the carotenoid biosynthesis pathway in three varieties of Cucurbita pepo with evident differences in fruit color. The transcriptional levels of the key genes involved in the carotenoid biosynthesis were higher in flower-, leaf-, and fruit skin tissues than flesh tissues. This correlated with higher concentration of carotenoid content in these tissues. The differential expression among the colored and white cultivars detected for some genes, such as LCYe, in combination with other regulatory mechanisms, could explain the large differences found in terms of carotenoid content among the three varieties. These results are a first step to elucidate carotenogenesis in C. pepo and demonstrate that, in general, regulation of the pathway genes is a critical factor that determines the accumulation of these compounds. © 2013 American Chemical Society.Peer Reviewe

Voxel Selection in MRI through Bagging and Conformal Analysis: Application to Detection of Obsessive Compulsive Disorder

In this work we apply a multivariate feature selection method based on bagging linear SVMs to construct a classifier able to differentiate among control subjects and patients with obsessive compulsive disorder (OCD). Our method selects sets of voxels that are relevant for the detection of the disease. The voxel selection is completed with a conformal analysis based refinement that controls over fitting and dramatically reduces the test error rate of the final classifier. Furthermore, the resulting discrimination pattern is organized in regions that show great agreement with those found by traditional methods used in OCD problems, achieving cleaner and more accurate region maps. © 2012 IEEE