RESUMEN
BACKGROUND: Xeroderma pigmentosum (XP) patients present a high risk of developing skin cancer and other complications at an early age. This disease is characterized by mutations in the genes related to the DNA repair system. OBJECTIVES: To describe the clinical and molecular findings in a cohort of 32 Brazilian individuals who received a clinical diagnosis of XP. METHODS: Twenty-seven families were screened for germline variants in eight XP-related genes. RESULTS: All patients (N = 32) were diagnosed with bi-allelic germline pathogenic or potentially pathogenic variants, including nine variants previously undescribed. The c.2251-1G>C XPC pathogenic variant, reported as the founder mutation in Comorian and Pakistani patients, was observed in 15 cases in homozygous or compound heterozygous. Seven homozygous patients for POLH/XPV variants developed their symptoms by an average age of 7.7 years. ERCC2/XPD, DDB2/XPE and ERCC5/XPG variants were found in a few patients. Aside from melanoma and non-melanoma skin tumours, a set of patients developed skin sebaceous carcinoma, leiomyosarcoma, angiosarcoma, mucoepidermoid carcinoma, gastric adenocarcinoma and serous ovarian carcinoma. CONCLUSIONS: We reported a high frequency of XPC variants in 32 XP Brazilian patients. Nine new variants in XP-related genes, unexpected non-skin cancer lesions and an anticipation of the clinical manifestation in POLH/XPV cases were also described.
Asunto(s)
Xerodermia Pigmentosa , Brasil , Niño , Reparación del ADN , Mutación de Línea Germinal , Homocigoto , Humanos , Mutación , Xerodermia Pigmentosa/genética , Proteína de la Xerodermia Pigmentosa del Grupo D/genéticaRESUMEN
OBJECTIVE: To analyze the relationship between therapeutic (weight-adjusted) dose of bemiparin and anti-Xa activity in patients with venous thromboembolism (VTE) and cancer in comparison with a cohort of patients with VTE without cancer, and its relationship with outcomes. MATERIALS AND METHODS: This is a prospective cohort study that comprised a cohort of patients with cancer-associated VTE and a cohort of non-cancer patients with VTE, all of them treated with bemiparin. The ethics committee approved the study and informed consent was obtained from the patients. RESULTS: One hundred patients were included (52 with cancer and 48 without cancer), with a median follow-up of 9.8 months. Mean anti-Xa activity was 0.89 (± 0.33) UI/mL in oncological patients and 0.83 (± 0.30) UI/mL in non-cancer patients (mean difference - 0.05 95% CI - 0.18; 0.06). A multiple linear regression model showed that anti-Xa peak was associated with the dose/kg independently of possible confounding variables (presence of cancer, age, sex and eGFR-estimated Glomerular Filtration Rate), in a way that for every 1 UI of dose/kg increase, the anti-Xa peak activity increased 0.006 UI/mL (95% CI 0.003; 0.009) (p < 0.001). The predictive capacity of anti-Xa peak in the oncology cohort showed an area under the ROC curve of 0.46 (95% CI 0.24-0.68), 0.70 (95% CI 0.49-0.91) and 0.74 (95% CI 0.44-0.94) for death, first bleeding and recurrence of VTE, respectively, and none was statistically significant. CONCLUSION: In patients with venous thromboembolism treated with bemiparin, anti-Xa levels were not influenced by the presence of cancer.
Asunto(s)
Anticoagulantes/uso terapéutico , Inhibidores del Factor Xa/sangre , Heparina de Bajo-Peso-Molecular/uso terapéutico , Neoplasias/complicaciones , Tromboembolia Venosa/sangre , Anciano , Anticoagulantes/efectos adversos , Femenino , Hemorragia/inducido químicamente , Heparina de Bajo-Peso-Molecular/efectos adversos , Humanos , Modelos Lineales , Masculino , Neoplasias/sangre , Estudios Prospectivos , Insuficiencia Renal/diagnóstico , Tromboembolia Venosa/tratamiento farmacológico , Tromboembolia Venosa/etiologíaRESUMEN
Los meningiomas son los tumores intracraneales benignos más frecuentes en adultos, suponen el 20% de todos los tumores cerebrales. Solamente el 1-4% presentan cambios quísticos. La relación entre el componente quístico y sólido de los meningiomas mixtos fue utilizada por varios autores para crear hasta cuatro clasificaciones diferentes de este subtipo de meningiomas. El diagnóstico mediante TC resulta muy complejo ante este subtipo de meningiomas por la dificultad de discernir entre diferentes lesiones intracraneales que también presentan componente quístico asociado. Presentamos el caso de un paciente varón de 49 años que presenta episodios de amaurosis fugax. En la TC se identificó una lesión sólida en localización frontotemporal izquierda, con gran componente quístico que producía importante efecto de masa y desplazaba la línea media. Se completó estudio mediante RM y arteriografía y finalmente fue diagnosticado como meningioma mixto, que tras embolización tumoral desde arteria meníngea media, se resecó quirúrgicamente.
Meningiomas are the most common benign intracranial tumors in adults, accounting for 20% of all brain tumors. Only 1-4% have cystic changes. The relationship between the cystic and solid component of mixed meningiomas was used by several authors to create up to four different classifications of this subtype of meningiomas. CT diagnosis is challenging in this subtype of meningiomas because of the difficulty of distinguishing between different intracranial lesions that also have associated cystic component. We present the case of a 49-year-old male patient with episodes of amaurosis fugax. On CT, a solid lesion was identified in the left frontotemporal region, with a large cystic component that produced an important mass effect and midline displacement. A study was completed by MR and arteriography and finally was diagnosed as mixed meningioma, which after tumor embolization via the middle meningeal artery, was surgically resected.
Asunto(s)
Humanos , Masculino , Persona de Mediana Edad , Tomografía Computarizada por Rayos X/métodos , Neoplasias Meníngeas/diagnóstico por imagen , Meningioma/diagnóstico por imagen , Espectroscopía de Resonancia Magnética , Meningioma/epidemiologíaRESUMEN
Neobenedenia melleni is a monogenean parasite that causes significant mortality and economic losses in fish aquaculture. Changes in the antigenic composition of this parasite occur during its developmental stages. In this study, we evaluated humoral parameters in serum and transcriptional immune responses of yellowtail naturally infected with N. melleni. In addition, in vitro assays were performed to study the stimulatory effects of antigens from larvae and adults on spleen leucocytes from non-infected fish at 6 and 24 h post-stimulation. The results showed enhanced total protein, myeloperoxidase and antiprotease activities in N. melleni-infected fish compared with non-infected ones. The induction of Toll-like receptors (TLRs) and pro-inflammatory cytokines in spleen leucocytes during natural infection with N. melleni suggests that these immune-related genes play an important role in the initiation of the immune defence mechanism for controlling parasite infection. Interestingly, the magnitude of in vitro responses of spleen leucocytes was dependent on the parasitic stage. An important stimulation of gene expression by adult antigens on spleen leucocytes was observed. Differential expression patterns of TLRs and target cytokines in yellowtail leucocytes in both in vivo and in vitro studies suggest that the quality of yellowtail immune response is conditioned by N. melleni development.
Asunto(s)
Antígenos Helmínticos/inmunología , Enfermedades de los Peces/inmunología , Inmunidad Humoral , Inmunidad Innata , Perciformes , Trematodos/inmunología , Infecciones por Trematodos/veterinaria , Animales , Citocinas/genética , Citocinas/metabolismo , Enfermedades de los Peces/parasitología , Proteínas de Peces/genética , Proteínas de Peces/metabolismo , Larva/genética , Larva/inmunología , Leucocitos/metabolismo , Bazo/metabolismo , Receptores Toll-Like/genética , Receptores Toll-Like/metabolismo , Trematodos/crecimiento & desarrollo , Infecciones por Trematodos/inmunología , Infecciones por Trematodos/parasitologíaRESUMEN
Rheumatoid arthritis (RA) is the prototype of the rheumatic diseases worldwide. Methotrexate (MTX) is the drug of first choice in the treatment of this disease due to its immunosuppressant effect. However, side events are present in 30% of the patients. The C677T and A1298C polymorphisms of the methylenetetrahydrofolate reductase (MTHFR) gene are involved in the metabolism of MTX. Earlier studies reported an association between these polymorphisms and elevation of hepatic enzymes. We analyzed the frequencies of both polymorphisms and the presence of transaminasemia in 70 Mexican patients with rheumatic arthritis treated with MTX. The 19% (13/70) of patients had an increase in the serum level of transaminases. The A1298C polymorphism was associated with elevation of transaminases (P=0.024). The identification of MTHFR genotypes for C677T and A1298C polymorphisms could lead clinicians to identify patients in risk of elevation of transaminases, and give them an individualized treatment, as is a goal of pharmacogenetics.
Asunto(s)
Antirreumáticos/efectos adversos , Artritis Reumatoide/tratamiento farmacológico , Artritis Reumatoide/genética , Metotrexato/efectos adversos , Metilenotetrahidrofolato Reductasa (NADPH2)/genética , Polimorfismo Genético , Transaminasas/sangre , Antirreumáticos/farmacocinética , Artritis Reumatoide/enzimología , Artritis Reumatoide/etnología , Estudios de Casos y Controles , Distribución de Chi-Cuadrado , Frecuencia de los Genes , Genotipo , Humanos , Metotrexato/farmacocinética , Metilenotetrahidrofolato Reductasa (NADPH2)/metabolismo , México/epidemiología , Oportunidad Relativa , Farmacogenética , Fenotipo , Medicina de Precisión , Medición de Riesgo , Factores de Riesgo , Resultado del Tratamiento , Regulación hacia ArribaRESUMEN
Larval midgut proteins of hematophagous parasites contain strong antigens that can be used for host immunization. This concept has been applied for immunization of Pelibuey sheep against Oestrus ovis L. (Diptera: Oestridae). The aim of this study was to examine the effect of immunization on larval establishment (LE) and development. Immunized lambs (I, n = 6) received two injections of crude gut membrane protein extracts (GMPE) from third instar larvae with Freund's incomplete adjuvant (FIA) on days 0 (Day of first immunization) and 21 (0.4 and 0.45 mg GMPE/lamb, respectively). The control group (C, n = 5) received physiological saline with FIA. Lambs were challenged with first instars on Day 29 (20 larvae) and Day 43 (25 larvae). Blood samples were collected biweekly and IgG titers were analyzed by ELISA. All lambs were slaughtered on Day 90 and number of larvae recovered, larval stage and larval weight were recorded at necropsy. No significant effect of immunization on LE (C = 28.9%; I = 31.0% P > 0.05) was observed. Antibody titers were higher in the immunized group on Day 28 (P < 0.05), but subsequently similar in both groups. Larval physiological age and weight were also significantly (P < 0.05) affected by immunization. Immunization of Pelibuey lambs with GMPE did not affect LE but did delay O. ovis larval development.
Asunto(s)
Dípteros , Inmunización/veterinaria , Miasis/veterinaria , Enfermedades de las Ovejas/prevención & control , Animales , Dípteros/crecimiento & desarrollo , Dípteros/inmunología , Ensayo de Inmunoadsorción Enzimática/métodos , Ensayo de Inmunoadsorción Enzimática/veterinaria , Adyuvante de Freund , Inmunoglobulina G/sangre , Larva/crecimiento & desarrollo , Larva/inmunología , Peso Molecular , Miasis/prevención & control , OvinosRESUMEN
Since haematological variables can be used to assess the health state in cultured fish, a haematological characterization of clinically healthy Oreochromis aureus was done to establish the reference indices of this species. Fish were subjected to different stressed conditions (bacterial infection, nitrite intoxication, malachite green overdose) to study the changes in the haematological indices and its relation with the health condition. This species showed microcytic anaemia under experimental bacterial infection by Corynebacterium sp.; anaemia, neutrophilia and erythrocytes deformation following nitrite intoxication and medication overdose with malachite green.
Asunto(s)
Cíclidos/sangre , Infecciones por Corynebacterium/veterinaria , Enfermedades de los Peces/sangre , Nitritos/toxicidad , Colorantes de Rosanilina/toxicidad , Anemia/inducido químicamente , Animales , Infecciones por Corynebacterium/sangre , Eritrocitos Anormales/efectos de los fármacos , Pruebas Hematológicas , Neutrófilos/metabolismoRESUMEN
One hundred and thirteen individuals were PCR-typed for nine STR loci with the AmpFlSTR Profiler Plus PCR amplification kit, including the following autosomal STRs: D3S1358, vWA, FGA, D8S1179, D21S11, D18S51, D5S818, D13S317 and D7S820. Allele frequencies for each STR were estimated, and they were compared to other populations. Genotype distribution by locus and by two-loci combination was in agreement with Hardy-Weinberg expectations for all nine STRs. For this region of Mexico, the combined probability of exclusion (PE) and power of discrimination (PD) were estimated: PE=99.964% and PD>99.999%.
Asunto(s)
Alelos , Genética de Población , Secuencias Repetidas en Tándem , Medicina Legal/métodos , Humanos , MéxicoRESUMEN
La Glicoproteína P (Gp-P) es una proteína transmembranaria con función transportadora, que tiene por misión expeler xenobióticos lipofilíticos. Esta glicoproteína tiene una distribución específica y unos niveles de expresión variables en las células sanguíneas humanas encargadas de la defensa en el organismo. El objetivo de este trabajo fue comparar y establecer si son reproducibles los resultados obtenidos por dos laboratorios distintos en la valoración de la expresión de glicoproteína P (Gp-P) en muestras celulares con baja expresión de esta glicoproteína, como es el caso de las células de sangre periférica. Se valoraron las subpoblaciones celulares de ciento veinticinco (125) muestras de sangre periférica de individuos sanos por citometría de flujo e inmunofluorescencia directa, utilizando el anticuerpo monoclonal JSB-1 conjugando con isotiocianato de fluoresceína (FITC), en dos citómetros de flujo de la misma marca y modelo, situados en dos laboratorios diferentes: Departamento de Anatomía Patológica, Facultad de Medicina, Universidad de Granada y Centro de Transfusión Sanguínea de Granada, España. Los resultados demostraron que la mejor correlación estadística del porcentaje de Positividad (por ciento POS) fue observada en los linfositos (r=0.93), mientras que para la Incorporación Media Fluorescencia Específica (IMFE) se observó en los granulocitos (r=0.94). Los resultados obtenidos con la metodología empleada en la determinación de Gp-P por citometría de flujo e inmunofluorescencia directa, permiten concluir que es una técnica reproducible y fiable para la evaluación del fenómeno de Resistencia Multiple a Drogas (MDR) en las subpoblaciones celulares normales de sangre periférica
Asunto(s)
Humanos , Citometría de Flujo , Leucocitos , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/sangre , Recolección de Muestras de Sangre , Medicina , VenezuelaRESUMEN
The susceptibility of 7 d old veliger larvae of the scallops Argopecten ventricosus and Nodipecten subnodosus, the penshell Atrina maura, and the Pacific oyster Crassostrea gigas to a pathogenic strain of Vibrio alginolyticus was investigated by challenging the larvae with different bacterial concentrations in a semi-static assay. The results indicate that the larvae of the 2 scallop species are more susceptible to the V. alginolyticus strain than those of the oyster and the penshell. Signs of the disease were similar to bacillary necrosis described in previous work. Interspecies differences in susceptibility to pathogens are discussed.
Asunto(s)
Moluscos/microbiología , Vibrio/patogenicidad , Animales , Susceptibilidad a Enfermedades , Larva/microbiología , Especificidad de la Especie , Factores de TiempoRESUMEN
Craniopharyngioma is the most common childhood tumor and thought to arise from embryonic remnants of Rathke's pouch. The paucity of published data on the molecular basis of these tumors prompted us to examine 22 adamantinomatous craniopharyngiomas looking for genetic abnormalities. Using the X-linked polymorphic androgen receptor gene as a tool for X-chromosome inactivating analysis, we found that a subset of craniopharyngiomas are monoclonal and therefore are probably due to acquired somatic genetic defects. Thus, we investigated these tumours for mutations within three candidate genes, Gsalpha, Gi2alpha and patched (PTCH). Using single stranded conformational polymorphism (SSCP), denaturing gradient gel electrophoresis and direct sequencing, the presence of somatic mutations in these genes could not be demonstrated in any tumor. Our data indicate that a subset of craniopharyngiomas are monoclonal and the mutations in the PTCH, Gsalpha, and Gi2alpha contribute little if any to craniopharyngioma development.
Asunto(s)
Adenoma/genética , Neoplasias Encefálicas/genética , Craneofaringioma/genética , Subunidades alfa de la Proteína de Unión al GTP Gi-Go/genética , Subunidades alfa de la Proteína de Unión al GTP Gs/genética , Proteínas Proto-Oncogénicas/genética , Adenoma/patología , Anticuerpos Monoclonales , Neoplasias Encefálicas/patología , Craneofaringioma/patología , Cartilla de ADN , Exones/genética , Subunidad alfa de la Proteína de Unión al GTP Gi2 , Humanos , Mutación/genética , Polimorfismo Conformacional Retorcido-Simple , Desnaturalización Proteica , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
To support a risk assessment of manganese exposure in two communities living within a manganese mining district a cross-sectional study was performed on a sample of the adult population of long-term residents. One community was exposed to a point source from an ore primary refining plant. Manganese is an essential mineral for human life. It is also the fourth in importance for industrial metal making. Data were collected on socioeconomic living conditions, emission sources, environmental media concentrations (air, water, soil, dust, food), respiratory symptomatology, and a neuropsychological examination (Mini-Mental Screening test, the Hooper Visual Organization test, the Ardila-Ostroski, and others). We examined 73 subjects (52 women), most of low socioeconomic status. Environmental air concentrations were 2 to 3 times higher than those in other urban concentrations. Manganese blood concentrations ranged from 7.5 to 88 microg/L, with a median concentration of 15, the upper quartile starting at 20 microg/L; the upper 10% was above 25 microg/L. Lead and manganese were highly correlated; there was an inverse relation to hemoglobin. Reduced levels of plasma lipid peroxidation were associated with blood manganese. Using multivariate logistic regression, we identified B-Mn as increasing the risk of deficient cognitive performance 12 times (Mini-Mental score of less than 17).
Asunto(s)
Trastornos del Conocimiento/inducido químicamente , Exposición a Riesgos Ambientales , Manganeso/efectos adversos , Salud Pública , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Estudios Transversales , Femenino , Humanos , Industrias , Plomo/efectos adversos , Plomo/sangre , Peroxidación de Lípido , Masculino , Manganeso/sangre , México , Persona de Mediana Edad , Proyectos Piloto , Medición de Riesgo , Clase Social , Población UrbanaRESUMEN
Modifications of microbial genomes often require the use of the antibiotic-resistance (Anb(R))-encoding genes and other easily selectable markers. We have developed a set of such selectable markers (Cm(R), Km(R) and Gm(R)), which could easily be inserted into the genome and subsequently removed by using the Cre/loxP site-specific recombination system of bacteriophage P1. In this manner the same marker could be used more than once in the same background, while the resulting strain could or would remain Anb(R) marker-free. Three plasmids were constructed, each containing a cassette consisting of the Cm(R), Km(R), or Gm(R) gene flanked by two parallel loxP sites and two polylinkers (MCS). To test insertion and excision, cassettes were inserted into the lacZ or galE genes carried on an origamma/pir-dependent suicide plasmid, which contained a dominant Sm(R) gene. The cassettes were crossed into the E. coli genome by homologous recombination (allelic exchange), in a manner analogous to that described by Pósfai et al. [Nucl. Acids Res. 22 (1994) 2392-2398], selecting for the Cm(R), Km(R), or Gm(R), for the LacZ(-) or GalE(-) and for the Sm(S) phenotypes (the latter to assure allelic exchange rather than insertion of the entire plasmid). When required, after selecting the strain with the desired modification, the Cm(R), Km(R), or Gm(R) marker was excised by supplying the Cre function. Cre was provided by the thermosensitive plasmid pJW168, which was transformed into the Anb(R) host at 30 degrees C, and was subsequently eliminated at 42 degrees C. Thus the Anb(R) marker was removed, whereas the lacZ or galE gene remained interrupted by the retained loxP site.
Asunto(s)
Bacterias/genética , Farmacorresistencia Microbiana/genética , Escherichia coli/genética , Genoma Bacteriano , Proteínas Virales , Acetiltransferasas/genética , Bacterias/efectos de los fármacos , Cloranfenicol/farmacología , Cloranfenicol O-Acetiltransferasa/genética , ADN Recombinante , Resistencia a Múltiples Medicamentos/genética , Escherichia coli/efectos de los fármacos , Eliminación de Gen , Marcadores Genéticos , Vectores Genéticos , Gentamicinas/farmacología , Integrasas/genética , Kanamicina/farmacología , Kanamicina Quinasa/genética , Operón Lac/genética , Mutagénesis Insercional , Plásmidos/genética , UDPglucosa 4-Epimerasa/genéticaRESUMEN
The aprE gene of Bacillus subtilis encodes the major serine alkaline protease known as subtilisin. It is expressed during the transition state and transcribed by the sigma(A) form of the RNA polymerase (RNAP). In this work, we characterized the regulatory region of the aprE gene (rraprE) from B. subtilis. By computer analysis and site-directed mutagenesis, we localized the aprE promoter sequence 7 bp upstream from its transcription initiation site (TIS). We also characterized the static curvature properties of the rraprE DNA and found two different areas of DNA bending, within the first 400 bp upstream of its TIS. We postulate that these particular curved DNA regions could play a role in the interaction with some regulatory proteins and discuss possible implications related to aprE transcription regulation.
Asunto(s)
Bacillus subtilis/genética , Proteínas Bacterianas/genética , ADN Circular/química , Regulación Bacteriana de la Expresión Génica , Proteínas de Transporte de Membrana , Regiones Promotoras Genéticas/genética , Subtilisinas/genética , Bacillus subtilis/metabolismo , Proteínas Bacterianas/metabolismo , Simulación por Computador , ADN Bacteriano/análisis , ADN Bacteriano/química , ADN Bacteriano/genética , ADN Circular/análisis , ADN Circular/genética , Electroforesis en Gel de Poliacrilamida , Mutagénesis Sitio-Dirigida , Plásmidos/genética , Análisis de Secuencia de ADN , Factor sigma , Subtilisinas/metabolismo , beta-Galactosidasa/metabolismoRESUMEN
A pBRINT-Ts family of integrative vectors for Escherichia coli was constructed by using a temperature-sensitive replicon derived from pSC101, a region of homology to the lacZ gene, and various antibiotic resistance markers (kanamycin, chloramphenicol and gentamycin) for selection of the integrants. The gene or group of genes to be integrated can be inserted into a multiple cloning site, flanked by an antibiotic resistance marker and lacZ sequences. A simple and rapid procedure was developed for the selection of cells where allelic exchange had occurred. With this procedure, the efficiency of integration of around 10-3 was observed, using several E. coli strains. From colonies with an integrated pBRINT-Ts plasmid, we detected an average allelic exchange event frequency of 7.5%. As a test for this system, we integrated the amy gene that codes for the alpha-amylase from B. stearothermophilus, into the lacZ gene of E. coli W3110. Production of alpha-amylase was found to be proportional to copy number; at up to 10 copies per chromosome.
Asunto(s)
Cromosomas Bacterianos , Escherichia coli/genética , Genética Microbiana/métodos , Plásmidos/genética , beta-Galactosidasa/genética , Clonación Molecular/métodos , Estabilidad de Enzimas , Geobacillus stearothermophilus/enzimología , Geobacillus stearothermophilus/genética , Reacción en Cadena de la Polimerasa , Temperatura , Transducción Genética , alfa-Amilasas/genética , beta-Galactosidasa/metabolismoRESUMEN
AIMS: The clinicopathological and immunohistochemical features of the second case of placental site nodule (PSN) of extrauterine, tubal location are presented. METHODS AND RESULTS: The lesion was incidentally found in the right tube during a cesarean section and eventual tubal ligation in a 23-year-old women gesta 2 para 1, after an uneventful 39-week intrauterine pregnancy. Grossly, the right Fallopian tube had a 1 cm dilatation filled by necrotic material. Microscopically, the lumen of the Fallopian tube was effaced and replaced by a rim of pleomorphic intermediate trophoblastic (IT) cells with pseudoinvasive parietal features which were positive for human placental lactogen, placental alkaline phosphatase, epithelial membrane antigen and CAM5.2. The Ki67 index was 3%. CONCLUSION: Due to its bizarre microscopic appearance, this lesion should be included in the differential diagnosis with malignant conditions. Both origins from a previous subclinical extrauterine tubal pregnancy and a possible migration of IT from a uterine implantation are considered.
Asunto(s)
Neoplasias de las Trompas Uterinas/diagnóstico , Tumor Trofoblástico Localizado en la Placenta/diagnóstico , Adulto , Biomarcadores/análisis , Neoplasias de las Trompas Uterinas/química , Femenino , Humanos , Técnicas para Inmunoenzimas , Embarazo , Tumor Trofoblástico Localizado en la Placenta/químicaRESUMEN
We have cloned and characterized the pykA gene from Bacillus subtilis which codes for a pyruvate kinase (PK) enzyme. This gene has been located downstream a putative phosphofructokinase gene, suggesting that they are part of the same operon. The deduced amino acid sequence of this PK showed a strong similarity to other PKs from different sources; however, as it has been found in other bacilli, the B. subtilis pykA enzyme had an extra C-terminal sequence consisting of about 112 amino acid residues. This gene was insertionally inactivated at the chromosomal level, with an antibiotic resistance marker. The analysis of this mutation in wild type and pts- backgrounds, indicated that B. subtilis has no other pyruvate kinase activity capable of complementing the absence of PykA.
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Bacillus subtilis/genética , Genes Bacterianos , Piruvato Quinasa/genética , Secuencia de Aminoácidos , Bacillus subtilis/fisiología , Secuencia de Bases , Clonación Molecular , Biblioteca de Genes , Prueba de Complementación Genética , Datos de Secuencia Molecular , Mutagénesis Insercional , Filogenia , Piruvato Quinasa/biosíntesis , Proteínas Recombinantes de Fusión/biosíntesis , Alineación de Secuencia , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Especificidad de la Especie , Esporas BacterianasRESUMEN
Plasmid pBRINT is an efficient vector for chromosomal integration of cloned DNA into the lacZ gene of Escherichia coli [Balbás et al., Gene 136(1993) 211-213]. A family of related plasmids containing different antibiotic-resistance markers (CmR or GmR or KmR) and a larger multiple cloning site (MCS) has been constructed. This set of plasmids, whose integration efficiencies are as good as those obtained with the prototype plasmid pBRINT, constitutes a collection of tools that allow rapid and easy integration of cloned DNA, at the chromosomal level. Their functionality as integration vectors has been ascertained by integrating the Vitreoscilla sp. hemoglobin-encoding gene and the Photobacterium leiognathi lux genes. To evaluate the level of expression obtained after chromosomal integration, we constructed strains carrying one or two copies of the cat gene integrated in the chromosome, and compared their enzymatic activities with those obtained from a strain carrying cat on a multicopy plasmid.
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Cromosomas Bacterianos , ADN/genética , Escherichia coli/genética , Plásmidos , Secuencia de Bases , Cloranfenicol O-Acetiltransferasa/genética , Clonación Molecular , Datos de Secuencia Molecular , Transducción GenéticaRESUMEN
The initiation of sporulation in Bacillus subtilis is under control of the transcriptional factor Spo0A. Most Spo0A mutants fail to initiate the sporulation process and all the sporulation initiated processes such as the synthesis of subtilisin. However, the product of spo0A9V, one of the several spo0A mutants characterized, distinguishes itself in the fact that, while it appears to effectively repress abrB, it fails to activate the spoIIA operon. The aim of this study was to examine the effect of the spo0A9V mutation on aprE expression and we found that in different genetic backgrounds, the spo0A9V mutation has a negative effect on aprE::lacZ expression.