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1.
Mem. Inst. Oswaldo Cruz ; 117: e210157, 2022. graf
Article in English | LILACS-Express | LILACS | ID: biblio-1375918

ABSTRACT

BACKGROUND Leishmania parasites cause leishmaniasis that range from self-limiting cutaneous lesions to more serious forms of the disease. The search for potential drug targets focusing on biochemical and metabolic pathways revealed the sterol biosynthesis inhibitors (SBIs) as a promising approach. In this class of inhibitors is found ketoconazole, a classical inhibitor of 14α-methysterol 14-demethylase. OBJECTIVE The present study aimed to better understand the biological response of Leishmania (Leishmania) amazonensis promastigotes at the cellular level after ketoconazole treatment. METHODS Herein, techniques, such as fluorimetry, flow cytometry, fluorescence microscopy, electron and scanning microscopy were used to investigate the cellular structures and to identify organelles affected by ketoconazole treatment. FINDINGS The study demonstrated, for the first time, the effect of ketoconazole on mitochondrion functioning and its probable relationship to cell cycle and death on L. (L.) amazonensis promastigotes (IFLA/BR/67/PH8 strain). MAIN CONCLUSIONS Ketoconazole-induced mitochondrial damages led to hyperpolarisation of this single organelle and autophagic vacuoles formation, as a parasite survival strategy. These damages did not reflect directly on the parasite cell cycle, but drove the parasites to death, making them susceptible to ketoconazole treatment in in vitro models.

2.
J. pediatr. (Rio J.) ; 97(3): 280-286, May-June 2021. tab, graf
Article in English | LILACS | ID: biblio-1279331

ABSTRACT

Abstract Objectives To evaluate the impact of invasive mechanical ventilation associated with two serum inflammatory cytokines and clinical indicators, on the second day of life, as predictors of bronchopulmonary dysplasia in very low birth weight preterm infants. It was hypothesized that the use of invasive mechanical ventilation in the first hours of life is associated with biomarkers that may predict the chances of preterm infants to develop bronchopulmonary dysplasia. Methods Prospective cohort of 40 preterm infants with gestational age <34 weeks and birth weight <1500 g. The following were analyzed: clinical variables; types of ventilator support used (there is a higher occurrence of bronchopulmonary dysplasia when oxygen supplementation is performed by long periods of invasive mechanical ventilation); hospitalization time; quantification of two cytokines (granulocyte and macrophage colony stimulating factor [GM-CSF] and eotaxin) in blood between 36 and 48 h of life. The preterm infants were divided in two groups: with and without bronchopulmonary dysplasia. Results The GM-CSF levels presented a significantly higher value in the bronchopulmonary dysplasia group (p = 0.002), while eotaxin presented higher levels in the group without bronchopulmonary dysplasia (p = 0.02). The use of continuous invasive mechanical ventilation was associated with increased ratios between GM-CSF and eotaxin (100% sensitivity and 80% specificity; receiver operating characteristic area = 0.9013, CI = 0.7791-1.024, p < 0.0001). Conclusions The duration of invasive mechanical ventilation performed in the first 48 h of life in the very low birth weight infants is a significant clinical predictor of bronchopulmonary dysplasia. The use of continuous invasive mechanical ventilation was associated with increased ratios between GM-CSF and eotaxin, suggesting increased lung injury and consequent progression of the disease.


Subject(s)
Humans , Infant, Newborn , Infant , Bronchopulmonary Dysplasia/diagnosis , Respiration, Artificial , Infant, Premature , Biomarkers , Prospective Studies
3.
Ciênc. rural (Online) ; 51(8): e20200945, 2021. tab, graf
Article in English | LILACS-Express | LILACS, VETINDEX | ID: biblio-1278898

ABSTRACT

ABSTRACT: Bovine genital campylobacteriosis (BGC) is a venereal and subclinical disease that affects the fertility of cattle herds, and it is caused by Campylobacter fetus subsp. venerealis . This study selected peptides mimetic to the BGC-causing agent from a phage library. Phage display is a technique that applies bacteriophage libraries that reveal peptides fused to the viral capsid in biological selections against target proteins. Biopannings were performed for biological selection in the phage library using rabbit hyperimmune serum and C. fetus subsp. venerealis protein extract. Five selected heptapeptides were considered mimetic to Cfv-NCTC 10354 based on the results of bioinformatics analysis and assays with hyperimmune serum and cervicovaginal mucus obtained from heifers. ALASLPL and LSYLFPP were the most reactive peptides and considered promising as possible mimetic immunogens for C. fetus subsp. venerealis.


RESUMO: Campilobacteriose Genital Bovina (CGB) é uma doença venérea e subclínica que causa problemas reprodutivos em rebanhos, causada por Campylobacter fetus subsp. venerealis. Este trabalho teve como objetivo selecionar peptídeos miméticos ao agente da CGB de uma biblioteca de fagos. Phage display é uma técnica que aplica bibliotecas de bacteriófagos que expõem peptídeos fusionados ao capsídeo viral em seleções biológicas contra proteínas alvo. Biopannings foram realizados para seleção biológica na biblioteca de fagos por meio de soro hiperimune de coelho e extrato proteico de C. fetus subsp. venerealis. Cinco heptapeptídeos selecionados foram considerados miméticos para Cfv-NCTC 10354 a partir de análises de bioinformática e ensaios com soro hiperimune e muco cérvico-vaginal de novilhas. ALASLPL e LSYLFPP foram os peptídeos mais reativos e considerados promissores como possíveis imunógenos miméticos para C. fetus subsp. venerealis.

4.
Genet. mol. biol ; 40(1): 61-68, Jan.-Mar. 2017. graf
Article in English | LILACS | ID: biblio-892362

ABSTRACT

Abstract Stingless bees of the genus Melipona, have long been considered an enigmatic case among social insects for their mode of caste determination, where in addition to larval food type and quantity, the genotype also has a saying, as proposed over 50 years ago by Warwick E. Kerr. Several attempts have since tried to test his Mendelian two-loci/two-alleles segregation hypothesis, but only recently a single gene crucial for sex determination in bees was evidenced to be sex-specifically spliced and also caste-specifically expressed in a Melipona species. Since alternative splicing is frequently associated with epigenetic marks, and the epigenetic status plays a major role in setting the caste phenotype in the honey bee, we investigated here epigenetic chromatin modification in the stingless bee Melipona scutellaris. We used an ELISA-based methodology to quantify global methylation status and western blot assays to reveal histone modifications. The results evidenced DNA methylation/demethylation events in larvae and pupae, and significant differences in histone methylation and phosphorylation between newly emerged adult queens and workers. The epigenetic dynamics seen in this stingless bee species represent a new facet in the caste determination process in Melipona bees and suggest a possible mechanism that is likely to link a genotype component to the larval diet and adult social behavior of these bees.

5.
Rev. Soc. Bras. Med. Trop ; 49(4): 398-407, July-Aug. 2016. tab, graf
Article in English | LILACS | ID: lil-792794

ABSTRACT

Abstract: Visceral leishmaniasis (VL) is one of the most important tropical diseases worldwide. Although chemotherapy has been widely used to treat this disease, problems related to the development of parasite resistance and side effects associated with the compounds used have been noted. Hence, alternative approaches for VL control are desirable. Some methods, such as vector control and culling of infected dogs, are insufficiently effective, with the latter not ethically recommended. The development of vaccines to prevent VL is a feasible and desirable measure for disease control; for example, some vaccines designed to protect dogs against VL have recently been brought to market. These vaccines are based on the combination of parasite fractions or recombinant proteins with adjuvants that are able to induce cellular immune responses; however, their partial efficacy and the absence of a vaccine to protect against human leishmaniasis underline the need for characterization of new vaccine candidates. This review presents recent advances in control measures for VL based on vaccine development, describing extensively studied antigens, as well as new antigenic proteins recently identified using immuno-proteomic techniques.


Subject(s)
Humans , Animals , Dogs , Antibodies, Protozoan/immunology , Protozoan Vaccines/immunology , Leishmania/immunology , Leishmaniasis, Visceral/prevention & control , Antigens, Protozoan/immunology , Protozoan Proteins/immunology , Leishmania/classification
6.
Rev. Soc. Bras. Med. Trop ; 48(6): 739-745, Nov.-Dec. 2015. tab, graf
Article in English | LILACS | ID: lil-767834

ABSTRACT

Abstract: INTRODUCTION: Despite multidrug therapy, leprosy remains a public health issue. The intradermal Bacillus Calmette-Guérin (BCG) vaccine, Mitsuda test (lepromin skin test), and anti-phenolic glycolipid I (PGL-I) serology are widely used in leprosy studies and have shown great epidemiological value. METHODS: This longitudinal study evaluated the relative risks and benefits of these three tools by comparing results observed in household contacts (HHCs) of leprosy patients who developed leprosy with those of HHCs who did not in a population of 2,992 individuals monitored during a 10-year period. RESULTS : Seventy-five (2.5%) new leprosy cases were diagnosed, including 28 (0.9%) co-prevalent cases. Therefore, for the risk-benefit assessment, 47 (1.6%) HHCs were considered as truly diagnosed during follow-up. The comparison between healthy and affected contacts demonstrated that not only did BCG vaccination increase protection, but boosters also increased to 95% relative risk (RR) reduction when results for having two or more scars were compared with having no scars [RR, 0.0459; 95% confidence interval (CI), 0.006-0.338]. Similarly, Mitsuda reactions >7mm in induration presented 7-fold greater protection against disease development compared to reactions of 0-3mm (RR, 0.1446; 95% CI, 0.0566-0.3696). In contrast, anti-PGL-I ELISA seropositivity indicated a 5-fold RR increase for disease outcome (RR, 5.688; 95% CI, 3.2412-9.9824). The combined effect of no BCG scars, Mitsuda reaction of <7mm, and seropositivity to anti-PGL-I increased the risk for leprosy onset 8-fold (RR, 8.109; 95% CI, 5.1167-12.8511). CONCLUSIONS: The adoption of these combined assays may impose measures for leprosy control strategies.


Subject(s)
Adolescent , Adult , Aged , Child , Child, Preschool , Female , Humans , Infant , Infant, Newborn , Male , Middle Aged , Young Adult , Antigens, Bacterial/immunology , BCG Vaccine/immunology , Contact Tracing/statistics & numerical data , Glycolipids/immunology , Leprosy/immunology , Enzyme-Linked Immunosorbent Assay , Longitudinal Studies , Lepromin/immunology , Leprosy/prevention & control , Leprosy/transmission , Risk Assessment
7.
Rev. Soc. Bras. Med. Trop ; 48(4): 370-379, July-Aug. 2015. ilus
Article in English | LILACS | ID: lil-755961

ABSTRACT

Abstract

Phage display is a high-throughput subtractive proteomic technology used for the generation and screening of large peptide and antibody libraries. It is based on the selection of phage-fused surface-exposed peptides that recognize specific ligands and demonstrate desired functionality for diagnostic and therapeutic purposes. Phage display has provided unmatched tools for controlling viral, bacterial, fungal, and parasitic infections, and allowed identification of new therapeutic targets to treat cancer, metabolic diseases, and other chronic conditions. This review presents recent advancements in serodiagnostics and prevention of leishmaniasis -an important tropical parasitic disease- achieved using phage display for the identification of novel antigens with improved sensitivity and specificity. Our focus is on theranostics of visceral leishmaniasis with the aim to develop biomarker candidates exhibiting both diagnostic and therapeutic potential to fight this important, yet neglected, tropical disease.

.


Subject(s)
Animals , Humans , Mice , Biomarkers , Cell Surface Display Techniques/methods , Leishmaniasis/diagnosis , Leishmaniasis/therapy , Vaccination , Biotechnology , Drug Discovery/methods , Genetic Techniques , Immunotherapy/methods , Leishmaniasis/immunology , Mice, Inbred BALB C
8.
Mem. Inst. Oswaldo Cruz ; 107(supl.1): 55-59, Dec. 2012. graf, tab
Article in English | LILACS | ID: lil-659741

ABSTRACT

Leprosy transmission still occurs despite the availability of highly effective treatment. The next step towards successfully eliminating leprosy is interrupting the chain of transmission of the aetiological agent, Mycobacterium leprae. In this investigation, we provide evidence that household contacts (HHCs) of leprosy patients might not only have subclinical infections, but may also be actively involved in bacilli transmission. We studied 444 patients and 1,352 contacts using anti-phenolic glycolipid-I (PGL-I) serology and quantitative polymerase chain reaction (qPCR) to test for M. leprae DNA in nasal swabs. We classified the patients according to the clinical form of their disease and the contacts according to the characteristics of their index case. Overall, 63.3% and 34.2% of patients tested positive by ELISA and PCR, respectively. For HHCs, 13.3% had a positive ELISA test result and 4.7% had a positive PCR test result. The presence of circulating anti-PGL-I among healthy contacts (with or without a positive PCR test result from nasal swabs) was considered to indicate a subclinical infection. DNA detected in nasal swabs also indicates the presence of bacilli at the site of transmission and bacterial entrance. We suggest that the concomitant use of both assays may allow us to detect subclinical infection in HHCs and to identify possible bacilli carriers who may transmit and disseminate disease in endemic regions. Chemoprophylaxis of these contacts is suggested.


Subject(s)
Humans , Antigens, Bacterial/blood , Family Characteristics , Glycolipids/blood , Leprosy/transmission , Mycobacterium leprae , Asymptomatic Infections , Antibodies, Bacterial/blood , Carrier State , DNA, Bacterial/analysis , Leprosy/diagnosis , Leprosy/epidemiology , Mycobacterium leprae/genetics , Mycobacterium leprae/immunology , Nasal Mucosa/microbiology , Polymerase Chain Reaction , Prevalence
9.
Rev. bras. parasitol. vet ; 18(1): 39-41, Mar. 2009. tab
Article in English | LILACS | ID: lil-606763

ABSTRACT

Phage display techniques have been widely employed to map epitope structures which have served as the basis for developing molecular vaccines. We have applied this technique to map specific epitopes of Rhipicephalus (Boophilus) microplus. In the present study, we have identified the potential immunogens using a process in which the selected phage clones were analyzed through bioinformatics, prior to final field tests. The present study demonstrates the feasibility of identifying important R. (B.) microplus phagotopes for vaccine development through screening of phage-displayed random peptide libraries and bioinformatics tools.


Técnicas de phage display têm sido amplamente empregadas para o mapeamento de epítopos os quais tem servido como base para o desenvolvimento de vacinas moleculares. Esta técnica foi aplicada no mapeamento de epítopos do Rhipicephalus (Boophilus) microplus. Neste estudo, potenciais imunógenos foram identificados pela adoção de um processo em que os clones de fagos foram analisados por bioinformática, previamente à realização dos testes. Os resultados demonstraram a possibilidade da identificação de importantes mimetopos do R. (B.) microplus para o desenvolvimento de vacinas através da seleção de bibliotecas de phage display associada à análise de bioinformática.


Subject(s)
Animals , Peptide Library , Ticks/classification
10.
Appl. cancer res ; 28(4): 153-160, Oct.-Dec. 2008. ilus, tab
Article in English | LILACS, Inca | ID: lil-519876

ABSTRACT

Objectives: The present work evaluates variations of polymorphic [CAG]n repeats present at exon 1 of the AR gene, as well as relative levels of its transcript, in order to investigate associations of these factors with prostatic tumor genesis in the Brazilian male population. Methods: Genomic DNA was extracted from blood samples from patients with prostate cancer (PCa), benign prostatic hyperplasia (BPH), and from a group of young Brazilian males to determine the number of [CAG]n repeats amplified by PCR. Mutation analysis in this amplified fragment was carried out using the LIS-SSCP technique. Total RNA was extracted from prostatic tissue to evaluate the AR gene transcript levels using semi-quantitative multiplex RT-PCR. Results: CAG length varied from 14 to 30, with an average of 21 repeats for PCa and the male group and 20 for the BPH group. No significant difference was found for [CAG]n polymorphism among the analyzed groups and there was no sporadic change in the amplified portion of the AR gene, nor loss of [CAG]n repeats, demonstrating that these do not contribute to the cancer occurrence. Nevertheless, the positive association between short alleles and TNM pT3 staging may indicate that CAG repeats is associated to PCa progression. The transcriptional levels were significantly increased in PCa than in BPH and were associated with serum PSA levels of 5-10 ng/mL. As diagnostic clinical parameter, the levels of AR gene presented 17-fold higher chance for PCa occurrence, 60% of sensibility and 95% of specificity. Conclusion: The data suggest that the highly miscegenated Brazilian male population presents a high frequency of [CAG]n short repeats, which may be associated with the PCa progression, while AR mRNA levels seems to be a good indicator of the incidence of this pathology, being useful in clinical practice for distinguishing patients with PCa from those with BPH.


Subject(s)
Humans , Gene Expression , Prostatic Hyperplasia , Prostatic Neoplasms , Prostatic Neoplasms/diagnosis , Prostatic Neoplasms/pathology
11.
Genet. mol. biol ; 29(2): 273-277, 2006. tab
Article in English | LILACS | ID: lil-432697

ABSTRACT

Estrogen has an important function in swine reproduction and growth. A Pvu II restriction enzyme polymorphism has been proven to be an important genetic variation in the estrogen receptor gene (ESR) and may be considered as a candidate gene for use in pig production but there is no data regarding the prevalence of this polymorphism in the Brazilian pig population. We used DNA samples from the following three purebred pig breeds: Large White (336 females and 26 males), Landrace (304 females and 27 males) and Pietrain (125 females and 11 males). The ESR genotyping was performed using PCR-RFLP. For each breed, genotypes for the ESR gene were compared independently for expected progeny differences (EPD) in litter size (LS), average daily weight gain (DWG) (g/day) and back fat thickness (BT) as measured in mm by ultrasound. In the Large White breed, but not the other breeds, the ESR genotype was significantly (p < 0.05) associated to LS, DWG and BT. Large Whites genotyped as AA or AB had higher EPD values for the LS and BT traits compared to BB Large Whites, while AA Large Whites had higher DWG EPD values than BB Large Whites. Our results for the Large White population showed that the A allele has a beneficial effect on LS, DWG and BT expected progeny differences.


Subject(s)
Animals , Receptors, Estrogen/genetics , Swine/genetics , Genetic Variation , Polymorphism, Genetic , Polymorphism, Restriction Fragment Length , Reproduction
12.
Rev. Assoc. Med. Bras. (1992) ; 51(3): 164-169, maio-jun. 2005. tab
Article in Portuguese | LILACS | ID: lil-411189

ABSTRACT

OBJETIVO: Estudar o efeito do polimorfismo M235T do gene do angiotensinogênio na doença arterial coronariana e na sua gravidade em pacientes com e sem infarto agudo do miocárdio. MÉTODOS: Estudo transversal com 305 indivíduos de raça branca que foram alocados em 2 grupos. O primeiro com 201 pacientes com doença arterial coronariana comprovada pela angiografia (lesão obstrutiva > 50 por cento), sendo 110 com infarto agudo do miocárdio e 91 sem infarto. O segundo, 104 indivíduos controles com artérias coronárias normais. O polimorfismo M235T do angiotensinogênio foi analisado através da genotipagem pela reação em cadeia da polimerase seguida da digestão pela enzima de restrição. RESULTADOS: A freqüência dos genótipos TT, MT e MM do angiotensinogênio não foi estatisticamente diferente entre os pacientes com doença arterial coronariana e os controles (chi2 = 0,123; p = 0,939) bem como nos grupos de infartados e não infartados (chi2 = 2,171; p = 0,338). O risco relativo de desenvolver doença arterial coronariana e de apresentar infarto analisado entre os genótipos TT vs MM, MT vs MM e TT+MT vs MM não foi significante. A análise da gravidade da doença aterosclerótica no grupo de pacientes com doença arterial coronariana mostrou não haver correlação com os genótipos; resultado semelhante foi encontrado na comparação entre os grupos com e sem infarto. CONCLUSÕES: Não há associação entre o polimorfismo M235T do gene do angiotensinogênio com a doença arterial coronariana, com a sua gravidade e nem com o infarto agudo do miocárdio.


Subject(s)
Female , Humans , Male , Middle Aged , Angiotensinogen/genetics , Coronary Artery Disease/genetics , Gene Frequency , Myocardial Infarction/genetics , Polymorphism, Genetic , Cross-Sectional Studies , Genotype , Phenotype
13.
Genet. mol. biol ; 28(1): 16-21, Jan.-Mar. 2005. ilus
Article in English | LILACS | ID: lil-399609

ABSTRACT

Fourty-six non-castrated, halothane-free, male Landrace pigs were genotyped by PCR-RFLP for the Rsa I polymorphism in the PIT1 gene and classified into AA and AB genotypes. Total RNA was extracted from the pituitaries and the relative quantities of growth hormone (GH) mRNA were determined by semi-quantitative RT-PCR. Pigs with the AB genotype had higher levels of GH mRNA than those with the AA genotype (p = 0.034; Kruskal-Wallis test). This result suggests that the Rsa I polymorphism may be involved in Pit-1 protein expression or function, which in turn may influence GH transcription and expression. Thus, the Rsa I PIT1 gene polymorphism in this pig line may be used as a molecular marker to identify higher GH expression and possibly select for carcass and performance traits affected by GH.


Subject(s)
Humans , Animals , Growth Hormone , Swine , Transcription Factors , Gene Expression , Pituitary Gland , Polymorphism, Genetic , RNA, Messenger
14.
Arq. bras. cardiol ; 83(5): 404-413, nov. 2004. ilus, tab
Article in English, Portuguese | LILACS | ID: lil-387189

ABSTRACT

RESUMO OBJETIVO:Avaliar a associação do polimorfismo A1166C do gene do receptor AT1 da angiotensina II (AT1R) com o infarto agudo do miocárdio e a severidade da doença arterial coronariana. MÉTODOS: Estudo prospectivo, transversal de 110 pacientes com infarto agudo do miocárdio submetidos à angiografia coronariana com lesão significante (> 50 por cento) avaliada por três critérios de severidade: número de vasos lesados, morfologia da placa aterosclerótica e escore de risco coronariano. Sem lesões coronarianas 104 indivíduos controles. O polimorfismo A1166C do gene do AT1R foi determinado pela reação em cadeia da polimerase no DNA dos leucócitos do sangue periférico. Os fatores de risco coronariano clássicos foram analisados em todos os indivíduos. RESULTADOS: Na estratificação dos genótipos em relação aos fatores de risco apenas o tabagismo teve predominância nos heterozigotos AC (p = 0,02). A freqüência dos genótipos nos pacientes infartados foi de AA = 54,5 por cento; AC = 35,5 por cento e CC = 10 por cento, sendo similar e não significativa em relação aos controles (p = 0,83). Não houve aumento do risco de infarto agudo do miocárdio nas comparações dos genótipos CC vs AA (OR = 1,35; IC-95 por cento = 0,50 - 3,59), AC vs AA (OR = 1,03; IC-95 por cento = 0,58 - 1,84 e AA+AC vs AA (OR = 1,33; IC-95 por cento = 0,51 - 3,45). Nenhum dos critérios de severidade teve associação significativa com os genótipos. CONCLUSAO: Os nossos resultados indicam não haver associação do polimorfismo A1166C do AT1R com o infarto agudo do miocárdio e nem com a severidade da doença arterial coronariana segundo nossos resultados.


Subject(s)
Humans , Male , Female , Coronary Artery Disease/genetics , Myocardial Infarction/genetics , Polymorphism, Genetic/genetics , Receptor, Angiotensin, Type 1/genetics , Case-Control Studies , Cross-Sectional Studies , DNA , Genetic Predisposition to Disease , Genotype , Logistic Models , Polymerase Chain Reaction , Prospective Studies , Risk Factors , Severity of Illness Index
15.
Biosci. j ; 19(2): 105-107, May-Aug. 2003. ilus
Article in Portuguese | LILACS | ID: lil-561903

ABSTRACT

A técnica Differential Display (DD), descrita primeiramente em 1992, promoveu um grande avanço nos estudos de expressão gênica em eucariotos, permitindo análises comparativas, isolamento e caracterização de novos genes relacionados aos mais variados processos biológicos. A larga aplicação do DD se deve, principalmente, ao fato de ser uma técnica rápida, fácil e necessitar de pequena quantidade de RNA total. Entretanto, o uso de radioisótopos em uma determinada etapa do DD restringe esse método a laboratórios que tenham áreas específicas para manipular esses reagentes e o torna inviável à grande maioria dos centros de pesquisa existentes no país. Assim, esse trabalho busca adaptar a coloração por nitrato de prata, não radioativa, à técnica do DD; utilizando como modelo biológico a espécie Apis mellifera (abelha africana). Os resultados têm sido promissores e mostram que a coloração por nitrato de prata é eficiente no DD e pode tornar a técnica mais difundida nos laboratórios nacionais, inclusive aplicada aos estudos de câncer.


Differential Display (DD), described in 1992, promoted a great progress in eukaryotic gene expression studies, allowing comparative analyses, isolation and characterization of new genes related to the most varied biological processes. The wide application of DD is due, mainly, to the fact of being a fast and easy technique and to the smalll amount of total RNA required. However, the radioisotope use in DD restricts this method to laboratories that have specific areas to manipulate those reagents and makes it unviable to the majority of the research centers in our country. Thus, this work adapted the silver nitrate staining (non radioactive) to the technique of DD using the Apis mellifera (africanizaed honeybee) as a biological model. The results were promising and the showed that the silver nitrate staining is efficiente in DD and it can turn the technique more applied the national laboratories.


Subject(s)
Animals , Bees , Bees , Gene Expression , Gene Expression Profiling , Polymerase Chain Reaction
17.
Braz. j. vet. res. anim. sci ; 40(5): 366-372, 2003. ilus, tab
Article in Portuguese | LILACS | ID: lil-359132

ABSTRACT

O aumento da produtividade e qualidade dos produtos animais vem se tornando de grande interesse econômico. A prolactina (PRL) é um hormônio essencial para o sucesso reprodutivo e seu receptor (RPRL) tem sido detectado em vários tecidos². O gene RPRL foi recentemente mapeado em suínos no cromossomo 16(6). Este trabalho teve como objetivo analisar a frequência genotípica do RPRL em três diferentes raças de suíno, Landrace, Large White e Pietrain e correlacionar os genótipos com características de interesse. Foram analisados um total de 124 animais. O DNA foi extraído de sangue total suíno e submetido a técnica de PCR-RFLP, para determinação do genótipo do gene do receptor da prolactina. As análises estatísticas mostraram que o genótipo RPRL teve efeito sobre peso médio diário na raça Landrace (p<0,0135). As médias de DEPGMD na raça Landrace também foram diferentes em relação ao genótipo (p< 0,0610), confirmando a análise dos dados reais de Ganho de Peso Médio Diário. Métodos de seleção assistida por marcadores, juntamente com métodos de seleção tradicional poderão ser utilizados para potencializar e acelerar o melhoramento de características de interesse econômico em suínos, onde o gene do receptor de prolactina (RPRL) poderá ser utilizado como um marcador molecular para o ganho de peso médio diário real e sua DEP.


Subject(s)
Animals , Genes , Prolactin , Receptors, Prolactin , Swine
18.
Genet. mol. biol ; 25(3): 277-279, Sept. 2002. tab
Article in English | LILACS | ID: lil-335766

ABSTRACT

Fixed bin frequencies for the VNTR loci D2S44, D4S139, D5S110, and D8S358 were determined in a Minas Gerais population sample. The data were generated by RFLP analysis of HaeIII-digested genomic DNA and chemiluminescent detection. The four VNTR loci have met Hardy-Weinberg equilibrium, and there was no association of alleles among VNTR loci. The frequency data can be used in forensic analyses and paternity tests to estimate the frequency of a DNA profile in the general Brazilian population


Subject(s)
Gene Frequency , Polymorphism, Restriction Fragment Length , Brazil
19.
Genet. mol. biol ; 25(3): 313-315, Sept. 2002. ilus, tab
Article in English | LILACS | ID: lil-335773

ABSTRACT

Molecular diagnostics are performed by using DNA from different body tissues. However, it is necessary to obtain genomic DNA of good quality. Due to the impossibility of collecting blood from slaughtered animals, DNA extraction from solid tissues is necessary. The objective of this study was to describe a protocol of DNA extraction from swine skin, adipose, brain, liver, kidney and muscle tissues. We obtained high molecular weight DNA of good quality, shown by agarose gel and amplification of two DNA fragments, 605bp and 891pb, by PCR. Spectrophotometric analysis of DNA concentration showed variation among the DNA from different tissues, with the liver and adipose tissues presenting the greatest and the smallest concentration, respectively. The described protocol has proven to be advantageous due to its simplicity, quickness, affordable reagents and absence of phenol, resulting in a high molecular weight DNA of good quality from several tissues


Subject(s)
Animals , DNA , Polymerase Chain Reaction , Swine
20.
Genet. mol. biol ; 25(1): 29-35, 2002. ilus, tab
Article in English | LILACS | ID: lil-324983

ABSTRACT

Genotype data of 477 animals of several swine races (Landrace - LD, Large White - LW, Pietrain - PI, LWXLDXPI, Piau, Monteiro, and unknown race) were obtained to determine the allele frequency of the obesity gene. Genotype data of 174 crossbred swine (LWXLDXPI) were also obtained, in order to assess its correlation with carcass evaluation data (lean meat percentage, backfat thickness at P2, loin eye area, adjacent fat area, total fat and meat). Finally, genotype data of 96 pure swine (Landrace, Large White and Pietrain) were collected, to establish its relation with meat quality (drip loss, meat color, texture analysis and intramuscular fat) and carcass evaluation data (lean meat percentage; ham, loin, shoulder and belly weights; and backfat thickness at P2). This work also aimed associating EPDs (expected progeny differences) for litter size, daily weight gain and backfat thickness with genotype data of 49 Large White males and 54 Landrace females. Genotyping was done on animal blood by PCR-RFLP, based on Stratil et al. (1997). Statistical analysis was done by using SAS software for variance analysis between genotypes and data for each cited class. For purebred animals, a mixed model was used, with sire within race as random effect. The allelic frequencies of alleles T and C were, respectively: 0.8142; 0.1857 (Landrace); 0.9125; 0.0875 (Large White); 0.9433; 0.0566 (Pietrain); 0.8333; 0.1666 (LWXLDXPI); 0.2500; 0.7500 (Piau); 0.8750; 0.1250 (Monteiro), and 0.8870; 0.1130 (unknown race). Since the highest allele C frequency occurred in Piau, we suggest that this allele could be associated with fat accumulation. In the Landrace race, a study was done separating the frequencies of 2 generations (great-grandfather and grandfather), and the differences confirmed by a Chi-square test, a higher frequency of allele C having been found in the grandparental generation. This suggests that this allele could be eliminated by selection from the great-grandparental generation, when the male grandparent is replaced by the great-grandparental generation. The obesity gene did not influence any of the carcass evaluation data from crossbred animals. In pure swine, where the only genotypes were TT and TC, it greatly influenced shoulder weight and meat texture, with the highest average in heterozygotes (shoulder: 4.07 vs. 3.93; texture: 2.62 vs. 1.82), suggesting better carcass quality and worse meat quality than in homozygotes


Subject(s)
Humans , Animals , Leptin , Obesity , Quantitative Trait, Heritable , Swine/genetics
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