Mariângela Cristofani

Identification of citrus hybrids through the combination of leaf apex morphology and SSR markers

Citrus breeding programs normally face several biological obstacles such as apomixis and polyembryony that result in a cumbersome identification of hybrid seedlings. The main purpose of this work is to describe the combined use of visual selection based on the leaf apex morphology and SSR analysis to differentiate hybrid from nucellar seedlings derived from the cross between the ‘Murcott’ tangor [Citrus reticulata Blanco × Citrus sinensis (L.) Osb.] and ‘Pêra’ sweet orange [Citrus sinensis(L.) Osb.]. A new morphological variable named leaf apex morphometric index is also described as the quantitative basis of the visual selection. The efficiency of visual selection of hybrids was tested under two growth conditions, seedlings germinated in seedbeds and in plastic tubes. Putative hybrid seedlings were also confirmed through the analysis of simple sequence repeats (SSR). The visual selection of hybrid seedlings resulted in an increase of 87.2% (p < 0.01) and 202.2% (p <0.001) in the number of correctly identified hybrids when compared to the method of random picking of seedlings in seedbeds and plastic tubes,respectively. The results indicate that the combination of visual selection and SSR analysis for the identification of hybrids derived from the cross of polyembryonic citrus cultivars will improve the accuracy of the selection,save time, and reduce the costs involved in the use of molecular markers alone in citrus breeding programs.

Genetic linkage maps of Citrus sunki Hort. ex. Tan. and Poncirus trifoliata (L.) Raf. and mapping of citrus tristeza virus resistance gene

A linkage map for Citrus sunki Hort. ex. Tan. and Poncirus trifoliate (L.) Raf cv. Rubidoux has been developed exploiting the pseudo-testcross mapping strategy and RAPD markers. From controlled crosses 314 F1 zygotic progeny individuals were obtained. Two separate data sets were generated, one for each parent. Preliminary linkage analysis was carried out with a phase-unknown backcross model at a LOD > 4.0 and max. Θ = 0.30. A total of 203 arbitrary 10-mer primers and 5 arbitrary 8-mer primers were screened for segregating RAPD markers. Pseudo-testcrosss mating configurations were displayed by 78 primers where the marker was present in one parent, absent in the other and segregating in the progeny. These 78 primers were run on 80 individuals of the progeny amplifying a total of 169 RAPD markers. Linkage analysis revealed that 125 of these RAPD loci fell into 18 linkage groups (10 groups for ‘Sunki’ and 8 groups for ‘Rubidoux’) while 44 loci remained unassigned to any linkage group. The total length of the maps was 732.32 cM for ‘Sunki’ and 866.88 cM for ‘Rubidoux’ with the distance between markers ranging from 0 to 43.8 cM. Bulked segregant analysis (BSA) was used to identify potential resistance-associated RAPD fragment markers. Two RAPD fragments were found flanking the CTV gene (OPAV12_470 – 17.8 cM and OPN15_1630 – 23.1 cM).

An in silico analysis of the key genes involved in flavonoid biosynthesis in Citrus sinensis

Citrus species are known by their high content of phenolic compounds, including a wide range of flavonoids. In plants, these compounds are involved in protection against biotic and abiotic stresses, cell structure, UV protection, attraction of pollinators and seed dispersal. In humans, flavonoid consumption has been related to increasing overall health and fighting some important diseases. The goals of this study were to identify expressed sequence tags (EST) in Citrus sinensis (L.) Osbeck corresponding to genes involved in general phenylpropanoid biosynthesis and the key genes involved in the main flavonoids pathways (flavanones, flavones, flavonols, leucoanthocyanidins, anthocyanins and isoflavonoids). A thorough analysis of all related putative genes from the Citrus EST (CitEST) database revealed several interesting aspects associated to these pathways and brought novel information with promising usefulness for both basic and biotechnological applications.

Avaliação in vitro de genótipos de citros a Phytophthora parasitica

The objective of this work was to evaluate the effect on citrus (Citrus spp.) rootstocks to Phytophthora parasitica by in vitro inoculation. Seedlings were cultivated in MS media for 40 days, under 18 h of light of photoperiod and at 25 oC. Inoculation was performed by injecting an infested needle into seedling collars. Evaluation was made by the length of lesions in centimeters. The experimental design was random with 15 replicates of each genotype. Results indicate good correlation between in vitro and orchard reactions genotypes to P. parasitica.