Browsing by Author "Cascardo, J.C.M."

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    Characterization of microsatellites from cacao–Moniliophthora perniciosa interaction expressed sequence tags
    (2008) Lima, L.S.; Gramacho, K.P.; Gesteira, A.S.; Lopes, U.V.; Gaiotto, F.A.; Zaidan, H.A.; Pires, J.L.; Cascardo, J.C.M.; Micheli, F.
    Theobroma cacao L.–Moniliophthora perniciosa expressed sequence tags (ESTs) were converted into useful satellite markers for population analysis and genetic mapping. Forty-nine flanking primer pairs from TSH1188 (a resistant genotype) and Catongo (a susceptible genotype) ESTs were designed and screened for polymorphism analysis. Eleven were polymorphic, with an average of 3.81 alleles per locus and a total of 42 alleles. The satellite markers were tested on 21 cacao accessions and two bulked DNAs generated from 6 resistant and 6 susceptible plants from a segregating F2 (SCA6 9 ICS1) population for witches’ broom resistance. These results show that EST-derived microsatellites (short sequence repeats, SSRs) in Theobroma cacao have many potential applications in linkage mapping and the planning of crosses.
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    dsRNA-induced gene silencing in Moniliophthora perniciosa, the causal agent of witches’ broom disease of cacao
    (2009) Caribe dos Santos, A.C.; Sena, J.A.L.; Santos, S.C.; Dias, C.V.; Pirovani, C.P.; Pungartnik, C.; Valle, R.R.; Cascardo, J.C.M.; Vincentz, M.
    The genome sequence of the hemibiotrophic fungus Moniliophthora perniciosa revealed genes possibly participating in the RNAi machinery. Therefore, studies were performed in order to investigate the efficiency of gene silencing by dsRNA. We showed that the reporter gfp gene stably introduced into the fungus genome can be silenced by transfection of in vitro synthesized gfpdsRNA. In addition, successful dsRNA-induced silencing of endogenous genes coding for hydrophobins and a peroxiredoxin were also achieved. All genes showed a silencing efficiency ranging from 18% to 98% when compared to controls even 28 d after dsRNA treatment, suggesting systemic silencing. Reduction of GFP fluorescence, peroxidase activity levels and survival responses to H2O2 were consistent with the reduction of GFP and peroxidase mRNA levels, respectively. dsRNA transformation of M. perniciosa is shown here to efficiently promote genetic knockdown and can thus be used to assess gene function in this pathogen.