RESUMO
BACKGROUND: The purpose of this study was to test the effectiveness of a DNA repair protocol in improving genetic testing in compromised samples, frequently encountered in Forensic Medicine. METHODS: In order to stretch the experiment conditions to the limits, as far as quality of samples and DNA is concerned, we tried the repair protocol on ten ancient human teeth obtained from an equal number of skeletons from a burial site in Lerna, Middle Helladic Greece (2100-1700 BC). For these samples, sex was previously determined morphologically, serving as a reference to compare our molecular data with. The samples were analysed using the DNA amelogenin sex test assay prior and after DNA polymerase repair. For every individual, two molecular sex determinations were obtained by visualising PCR products on an agarose gel. RESULTS: DNA repair enabled genetic testing in these samples. Successful amplification of the amelogenin gene was obtained only from the repaired DNA in eight out of ten samples. Prior to the repair treatment, none of these samples yielded any PCR products, thus attesting to the authenticity of the amplified sequence. The concordance between morphological and molecular analysis was in reasonable agreement (71%). CONCLUSIONS: These results reveal the impact of the repair process in studying single copy genes from low quality DNA. This protocol could facilitate molecular analysis in compromised samples, encountered in forensic medicine, as well as enable genetic studies in ancient remnants.
RESUMO
A large scale microarray (20k MMC1) from the African malaria vector Anopheles gambiae was used to monitor gene expression in insecticide resistant and susceptible strains of the Asian mosquito Anopheles stephensi. Heterologous hybridization at slightly reduced stringency yielded approximately 7000 significant signals. Thirty-six putative genes were differentially transcribed between the pyrethroid-resistant (DUB-R) and the susceptible (BEECH) strains. The expression profiles of selected transcripts were verified by real-time PCR. A gene putatively involved in the thickening of the adult cuticle showed the most striking up-regulation in DUB-R. A more specialized microarray containing 231 An. gambiae genes putatively involved in insecticide detoxification was used to further analyse classical insecticide resistance genes. Three glutathione S-transferase (GST) transcripts, one esterase and a cytochrome P450 were up-regulated in the resistant strain, while two peroxidases were down-regulated.
Assuntos
Anopheles/genética , Etiquetas de Sequências Expressas , Perfilação da Expressão Gênica , Proteínas de Insetos/metabolismo , Resistência a Inseticidas/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Primers do DNA , Dados de Sequência Molecular , Análise de Sequência com Séries de Oligonucleotídeos , Análise de Sequência de DNA , Especificidade da EspécieRESUMO
Cytochrome P450s are a superfamily of haemoproteins, important in the metabolism of endogenous compounds and xenobiotics. As a first step to elucidating the role of this family in insecticide resistance in the malaria mosquito, Anopheles gambiae, we have cloned and mapped multiple P450 genes. Sixteen cDNAs encoding full-length P450s were cloned and physically mapped to the mosquito's polytene chromosomes. Fourteen of these encode putative CYP6 proteins and two encode P450s belonging to the CYP9 class. Eighteen new A. gambiae Cyp4 P450 genes were identified using degenerate PCR primers, cDNAs were detected for ten and in situ locations for thirteen members of this gene family.
Assuntos
Anopheles/enzimologia , Sistema Enzimático do Citocromo P-450/genética , Genes de Insetos , Insetos Vetores/enzimologia , Sequência de Aminoácidos , Animais , Anopheles/genética , Sequência de Bases , DNA Complementar , Insetos Vetores/genética , Malária , Dados de Sequência Molecular , Homologia de Sequência de AminoácidosRESUMO
PURPOSE: To study the effect of external beam irradiation on the morphometry of both angioplasted and nonangioplasted arteries in a hypercholesterolemic rabbit model. METHODS AND MATERIALS: Four groups of rabbit femoral arteries were studied: arteries (a) with no intervention, (b) irradiated with a 12-Gy x-ray dose, (c) treated with balloon angioplasty, and (d) dosed with 12 Gy 30 min after balloon angioplasty. RESULTS: External irradiation did not change vessel morphometry in nonangioplasted arteries. On the contrary, it induced neointimal formation and decreased luminal area, without causing any vessel remodeling in arteries treated with balloon angioplasty. CONCLUSION: External irradiation at 12 Gy given 30 min after angioplasty in the studied model accentuated the neointimal response to vascular injury, without causing any vessel remodeling.