Genotyping, serotyping and determination of mating-type of Cryptococcus neoformans clinical isolates from São Paulo State, Brazil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

THE USE OF AMPPD AS AN ALTERNATIVE SUBSTRATE FOR AP-MEDIATED DETECTION OF NONRADIOLABELED DNA PROBES IN EUCALYPTUS-SALIGNA

We present a non-radioactive alternative to Southern's (J. Mol. Biol. 98: 503-517, 1975) DNA-DNA hybridization technique. The use of AMPPD - Disodium 3-(4-Methoxyspiro {1,2-dioxetane-3,2'tricyclo[3.3.1.1(3,7)]decan}-4-yl)phyenyl phosphate as an alternative substrate for AP-mediated detection of digoxigenin-11 dUTP-labeled probes made possible the simple and nonhazardous reuse of blots. We used 0.8 % agarose gels containing 30 mug per lane of Eucalyptus saligna DNA, digested with Eco RI, electrophoresed and blotted on to nylon membranes (Hybond-N, Amersham, UK), using the Southern blotting procedure, and UV irradiated for one minute for DNA fixation. The hybridizations were carried out overnight with digoxigenin labeled random inserts of E. saligna DNA by using the Genius Kit (Boehringer Mannheim). Detection of the DNA-DNA hybrids was performed in the presence of 0.5% blocking agent and the substrates NBT/BCIP were replaced by 0.26 mM AMPPD in the final alkaline assay buffer (50 mul/cm2). After membrane incubation for five minutes at room temperature in a sealed plastic bag, the AMPPD solution was retrieved and stored at 4-degrees-C for reuse. A Kodak X-BRAF QA-S film was pressed firmly onto the bag containing the wet membrane, exposed for two to six hours and then developed. After use, the probes were stripped off and the blots reutilized, three times so far, with the same results.

Lack of reliability of nanotechnology in the of free plasma DNA in samples of patients with prostate cancer

Background: Several studies seek biological markers that give diagnostic and degree of tumor development. The aim of this study was to validate the determination of plasma DNA using nanotechnology (Nanovue™-NV) in samples of 80 patients with prostate cancer. Methods. Blood samples of 80 patients of the Urology Ambulatory of Faculdade de Medicina do ABC with prostate cancer confirmed by anatomical-pathology criteria were analyzed. DNA extraction was performed using a GFX TM kit (Amersham Pharmacia Biotech, Inc, USA) following the adapted protocol. Plasma was subjected to centrifugation. Results: There was a big difference between the first and the second value obtained by NanoVue Only two samples had no differences between duplicates. Maximum difference between duplicates was 38 μg/mL. Average variation between 51 samples was 10.29 μg/mL, although 21 samples had differences above this average. No correlation was observed between pDNA obtained by traditional spectrophotometry and by nanotechnology. Conclusion: Determination of plasma DNA by nanotechnology was not reproducible. © 2013 Moreno et al; licensee BioMed Central Ltd.

Caracterização da estrutura molecular e propriedades viscográficas de amidos de mandioca (Manihot esculenta Crantz) para aplicações industriais

Pós-graduação em Agronomia (Energia na Agricultura) - FCA

Avaliação da diversidade genética de populações de Pacu (Piaractus mesopotamicus Holmberg, 1887) do Pantanal Matogrossense com o uso de marcadores moleculares do tipo microssatélites

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Associação da mutação G1691A (fator V de Leiden) no gene do fator V da coagulação, da mutação G20210A no gene da protrombina e das mutações C677T e G1793A no gene da metilenotetrahidrofolato redutase com a doença arterial coronariana

Pós-graduação em Biociências e Biotecnologia Aplicadas à Farmácia - FCFAR