RESUMEN
AIMS: To give an updated perspective of interventions from additional data collected since our first review, conducted in 2008. BACKGROUND: This updated Cochrane Review incorporates new information from recent randomized controlled trials on culturally appropriate diabetes health education interventions. METHODS: An electronic literature search of six databases was repeated, with databases of ongoing trials checked and three journals hand-searched. Meta-analysis was carried out for sufficiently homogeneous outcomes, and common themes among trials were highlighted. RESULTS: A total of 22 new trials were added to the original 11. Meta-analysis of 28 trials containing suitable data showed significant improvements in glycaemic control (HbA1c ) and diabetes knowledge over a period of 24 months, after the delivery of culturally appropriate education to participants, compared with those receiving 'conventional' care. There were no consistent benefits over the control group in other selected outcome measures, and lack of data continued to make analysis of several outcome measures difficult. CONCLUSIONS: Research activity in this field has increased considerably over the past 6 years, with culturally appropriate diabetes education showing consistent benefits over conventional care in terms of glycaemic control and diabetes knowledge, sustained in the short- to mid-term. Further research is needed to determine the clinical significance of these improvements and their cost-effectiveness.
Asunto(s)
Asistencia Sanitaria Culturalmente Competente , Diabetes Mellitus Tipo 2/terapia , Medicina Basada en la Evidencia , Disparidades en el Estado de Salud , Salud de las Minorías , Educación del Paciente como Asunto , Terapia Combinada/tendencias , Asistencia Sanitaria Culturalmente Competente/tendencias , Países Desarrollados , Complicaciones de la Diabetes/etnología , Complicaciones de la Diabetes/prevención & control , Diabetes Mellitus Tipo 2/complicaciones , Diabetes Mellitus Tipo 2/epidemiología , Diabetes Mellitus Tipo 2/etnología , Conocimientos, Actitudes y Práctica en Salud/etnología , Política de Salud/tendencias , Transición de la Salud , Humanos , Hipoglucemia/prevención & control , Salud de las Minorías/educación , Salud de las Minorías/tendencias , Cooperación del Paciente/etnología , Educación del Paciente como Asunto/tendencias , Prevalencia , Ensayos Clínicos Controlados Aleatorios como Asunto , Reproducibilidad de los ResultadosRESUMEN
The removal of the mercaptan, 1-methyl-1-propanethiol, from aqueous solutions using a non-porous, electrically conducting carbon-based adsorbent (Nyex 1000) was investigated. The adsorption process was found to be rapid (equilibrium capacity achieved within 5 minutes) with low adsorptive capacity (of the order of 0.4 mg g(-1)) when compared with activated carbon. Electrochemical regeneration of the Nyex 1000 in a simple divided electrochemical cell within a sequential batch treatment unit restored 100% of the adsorbent's adsorptive capacity using treatment times as low as 20 minutes by passing a current of 0.5 A. The sorptive characteristics of a Nyex-water slurry were also modelled and investigated both in a bubble column and in a continuous adsorption-regeneration treatment system. It was demonstrated that the continuous removal-destruction system could achieve a step reduction in challenge gas concentration of approximately 75% for a period of 35 minutes with a current of 5 Amps. This was attributed to mass transfer enhanced by a combination of adsorption and chemical reaction with free chlorine species generated in the electrochemical process.
Asunto(s)
Compuestos de Sulfhidrilo/química , Adsorción , Contaminantes Atmosféricos/química , Contaminantes Atmosféricos/aislamiento & purificación , Carbón Orgánico/química , Electroquímica , Compuestos de Sulfhidrilo/aislamiento & purificación , Contaminantes Químicos del Agua/química , Contaminantes Químicos del Agua/aislamiento & purificaciónRESUMEN
Prolonged hypoxia, encountered in individuals suffering from various cardiorespiratory diseases, enhances the likelihood of subsequently developing Alzheimer's disease (AD). However, the underlying mechanisms are unknown, as are the mechanisms of neurodegeneration of amyloid beta peptides (AbetaPs), although the latter involves disruption of Ca2+ homeostasis. Here, immunohistochemistry demonstrated that hypoxia increased production of AbetaPs, an effect which was prevented by inhibition of either beta or gamma secretase, the enzymes required for liberation of AbetaP from its precursor protein. Whole-cell patch clamp recordings showed that hypoxia selectively increased functional expression of L-type Ca2+ channels. This was prevented by inhibition of either beta or gamma secretase, indicating that hypoxic channel up-regulation is dependent upon AbetaP formation. Our results indicate for the first time that hypoxia promotes AbetaP formation in central neurons, and show that this leads to abnormally high selective expression of L-type Ca2+ channels whose blockade has previously been shown to be neuroprotective in AD models. These findings provide a cellular basis for understanding the increased incidence of AD following prolonged hypoxia.
Asunto(s)
Péptidos beta-Amiloides/metabolismo , Canales de Calcio Tipo L/metabolismo , Cerebelo/metabolismo , Potenciales de la Membrana/fisiología , Neuronas/metabolismo , Oxígeno/metabolismo , Animales , Hipoxia de la Célula/fisiología , Células Cultivadas , RatasRESUMEN
Neurofibrillary tangles (NFTs) are the most common intraneuronal inclusion in the brains of patients with neurodegenerative diseases and have been implicated in mediating neuronal death and cognitive deficits. Here, we found that mice expressing a repressible human tau variant developed progressive age-related NFTs, neuronal loss, and behavioral impairments. After the suppression of transgenic tau, memory function recovered, and neuron numbers stabilized, but to our surprise, NFTs continued to accumulate. Thus, NFTs are not sufficient to cause cognitive decline or neuronal death in this model of tauopathy.
Asunto(s)
Encéfalo/metabolismo , Memoria , Enfermedades Neurodegenerativas/patología , Enfermedades Neurodegenerativas/fisiopatología , Ovillos Neurofibrilares/patología , Proteínas tau/metabolismo , Envejecimiento , Animales , Atrofia , Encéfalo/patología , Cognición , Progresión de la Enfermedad , Doxiciclina/farmacología , Hipocampo/metabolismo , Hipocampo/patología , Humanos , Aprendizaje por Laberinto , Ratones , Ratones Transgénicos , Enfermedades Neurodegenerativas/metabolismo , Ovillos Neurofibrilares/metabolismo , Plasticidad Neuronal , Neuronas/metabolismo , Neuronas/patología , Tamaño de los Órganos , Fosforilación , ARN Mensajero/genética , ARN Mensajero/metabolismo , Solubilidad , Proteínas tau/química , Proteínas tau/genéticaRESUMEN
As a normal consequence of aging, men experience a significant decline in androgen levels. Although the neural consequences of age-related androgen depletion remain unclear, recent evidence suggests a link between low androgen levels and the development of Alzheimer's disease (AD). Here, we test the hypothesis that androgens act as endogenous modulators of beta-amyloid protein (Abeta) levels. To investigate this possibility, brain and plasma levels of Abeta were measured in male rats with varying hormonal conditions. Depletion of endogenous sex steroid hormones via gonadectomy (GDX) resulted in increased brain levels of Abeta in comparison to gonadally intact male rats. This GDX-induced increase in Abeta levels was reversed by DHT supplementation, demonstrating a functional role for androgens in modulating brain levels of Abeta. These findings suggest that age-related androgen depletion may result in accumulation of Abeta in the male brain and thereby act as a risk factor for the development of AD.
Asunto(s)
Péptidos beta-Amiloides/metabolismo , Andrógenos/fisiología , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Péptidos beta-Amiloides/sangre , Andrógenos/administración & dosificación , Animales , Dihidrotestosterona/administración & dosificación , Implantes de Medicamentos , Estradiol/administración & dosificación , Terapia de Reemplazo de Hormonas , Masculino , Orquiectomía , Ratas , Ratas Sprague-Dawley , Receptores Androgénicos/metabolismo , Núcleos Septales/citología , Núcleos Septales/efectos de los fármacos , Núcleos Septales/metabolismoRESUMEN
Testosterone has been shown to have multiple beneficial effects on neuronal viability in developing and adult animals. Most often, testosterone promotes neural health indirectly via enzymatic conversion to estradiol by aromatase. Unclear is whether androgens can directly modulate vulnerability to neuronal insults in adult animals. We investigated this issue by modulating androgen status in rats prior to challenge with the excitotoxin kainate. Adult male rats were maintained in the following conditions: i) gonadectomized (GDX) to deplete endogenous androgens, ii) GDX+replacement with dihydrotestosterone (DHT) the active and non-aromatizable testosterone metabolite, iii) sham-GDX. Animals were then lesioned with kainate and surviving hippocampal neurons quantified. In the CA2/3 and hilar regions of the hippocampus, a modest lesion was observed in sham-GDX animals corresponding to approximately 25% cell loss in comparison to non-lesioned rats. The depletion of endogenous androgens by GDX significantly augmented lesion severity, consistent with the hypothesis that androgens are involved in maintaining cell viability. Importantly, DHT hormone replacement in GDX rats significantly attenuated kainate-induced neuron loss in CA2/3, suggesting direct androgen neuroprotection. These results demonstrate that androgens act as endogenous modulators of neuron viability, a function that may be compromised in aging men as a consequence of normal, age-related androgen depletion.
Asunto(s)
Andrógenos/farmacología , Dihidrotestosterona/farmacología , Agonistas de Aminoácidos Excitadores/toxicidad , Ácido Kaínico/toxicidad , Neuronas/efectos de los fármacos , Animales , Conducta Animal/efectos de los fármacos , Recuento de Células , Supervivencia Celular/efectos de los fármacos , Interacciones Farmacológicas , Hipocampo/citología , Hipocampo/efectos de los fármacos , Hipocampo/metabolismo , Inmunohistoquímica/métodos , Masculino , Neuronas/metabolismo , Orquiectomía/métodos , Fosfopiruvato Hidratasa/metabolismo , Ratas , Ratas Sprague-Dawley , Tiempo de Reacción/efectos de los fármacos , Receptores Androgénicos/metabolismo , Convulsiones/inducido químicamente , Convulsiones/prevención & control , Tabique del Cerebro/citología , Tabique del Cerebro/efectos de los fármacos , Tabique del Cerebro/metabolismoRESUMEN
The effects of amyloid beta protein on voltage-gated K(+) channel currents were studied using the whole-cell patch-clamp technique. The 1-40 amino acid form of amyloid beta protein was applied to primary cultures of rat cerebellar granule and cortical neurones for 24 h. Both the unaggregated and aggregated forms of the peptide, which have differing biological activities, were used. In cerebellar granule neurones, 24-h pre-incubation with 1 microM unaggregated amyloid beta protein resulted in a 60% increase in the 'A'-type component of K(+) current. Increased delayed rectifier activity was Cd(2+)-sensitive and was presumed to be secondary to an increase in voltage-gated Ca(2+) channel current activity. Unaggregated amyloid beta protein had no effect on any component of the K(+) channel current in cortical neurones. One micromolar of aggregated amyloid beta protein had no effect on K(+) channel current in either cell type but reduced cell survival within 24 h as measured using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling (TUNEL) assays. The unaggregated form of amyloid beta protein had no neurotoxic effects when applied to either neurone type for up to 72 h. These data indicate that the unaggregated, non-pathological form of amyloid beta protein causes changes in the ion channel function of neurones, possibly reflecting a physiological role for the peptide.
Asunto(s)
Péptidos beta-Amiloides/toxicidad , Activación del Canal Iónico/efectos de los fármacos , Neuronas/fisiología , Fragmentos de Péptidos/toxicidad , Canales de Potasio/fisiología , Enfermedad de Alzheimer/fisiopatología , Péptidos beta-Amiloides/química , Animales , Cloruro de Cadmio/farmacología , Células Cultivadas , Cerebelo/citología , Corteza Cerebral/citología , Potenciales de la Membrana/efectos de los fármacos , Neuronas/efectos de los fármacos , Técnicas de Placa-Clamp , Fragmentos de Péptidos/química , Potasio/metabolismo , Ratas , SolubilidadRESUMEN
Beta-amyloid protein is thought to underlie the neurodegeneration associated with Alzheimer's disease by inducing Ca(2+)-dependent apoptosis. Elevated neuronal expression of the proinflammatory cytokine interleukin-1beta is an additional feature of neurodegeneration, and in this study we demonstrate that interleukin-1beta modulates the effects of beta-amyloid on Ca(2+) homeostasis in the rat cortex. beta-Amyloid-(1-40) (1 microM) caused a significant increase in (45)Ca(2+) influx into rat cortical synaptosomes via activation of L- and N-type voltage-dependent Ca(2+) channels and also increased the amplitude of N- and P-type Ca(2+) channel currents recorded from cultured cortical neurons. In contrast, interleukin-1beta (5 ng/ml) reduced the (45)Ca(2+) influx into cortical synaptosomes and inhibited Ca(2+) channel activity in cultured cortical neurons. Furthermore, the stimulatory effects of beta-amyloid protein on Ca(2+) influx were blocked following exposure to interleukin-1beta, suggesting that interleukin-1beta may govern neuronal responses to beta-amyloid by regulating Ca(2+) homeostasis.
Asunto(s)
Péptidos beta-Amiloides/fisiología , Canales de Calcio Tipo L/fisiología , Calcio/metabolismo , Interleucina-1/fisiología , Neuronas/metabolismo , Fragmentos de Péptidos/fisiología , Sinaptosomas/metabolismo , Animales , Células Cultivadas , Corteza Cerebral/citología , Corteza Cerebral/metabolismo , Homeostasis , Activación del Canal Iónico , Transporte Iónico , RatasRESUMEN
We describe a new method of gene cloning by complementation of mutant alleles which obviates the need for construction of a gene library in a plasmid vector in vitro and its amplification in Escherichia coli. The method involves simultaneous transformation of mutant strains of the fungus Aspergillus nidulans with (i) fragmented chromosomal DNA from a donor species and (ii) DNA of a plasmid without a selectable marker gene, but with a fungal origin of DNA replication ('helper plasmid'). Transformant colonies appear as the result of the joining of chromosomal DNA fragments carrying the wild-type copies of the mutant allele with the helper plasmid. Joining may occur either by ligation (if the helper plasmid is in linear form) or recombination (if it is cccDNA). This event occurs with high efficiency in vivo, and generates an autonomously replicating plasmid cointegrate. Transformants containing Penicillium chrysogenum genomic DNA complementing A. nidulans niaD, nirA and argB mutations have been obtained. While some of these cointegrates were evidently rearranged or consisted only of unaltered replicating plasmid, in other cases plasmids could be recovered into E. coli and were subsequently shown to contain the selected gene. The utility of this "instant gene bank" technique is demonstrated here by the molecular cloning of the P. canescens trpC gene.
Asunto(s)
Aspergillus nidulans/genética , Clonación Molecular/métodos , ADN de Hongos , Penicillium/genética , Escherichia coli , Biblioteca de Genes , Prueba de Complementación Genética , Mutación , Plásmidos , Mapeo Restrictivo , Transformación GenéticaRESUMEN
Promoter activity was examined in the beta-lactam-producing fungus, Acremonium chrysogenum, by assessment of the properties of transformant isolates. Transformation was achieved using plasmid constructs specifying hygromycin B resistance (HyR) linked to the promoter elements of gpdA (the glucose-6-phosphate dehydrogenase-encoding gene of Aspergillus nidulans), and pcbC [the gene encoding the isopenicillin N synthetase (IPNS) enzyme of A. chrysogenum]. Transformation frequency, HyR levels, and Hy phosphotransferase (HPT) levels suggested that the transformants of constructs using the gpdA promoter showed a higher level of expression of the HyR gene than in transformants obtained using the pcbC promoter. The patterns of integration of the transforming DNA also differed in that pcbC promoter construct transformants appeared to have tandem repeats. All integrations of plasmid DNA occurred on a single chromosome which was different in four out of five transformants studied. Multiple copy transformants of constructs using the pcbC promoter did not show the regulated pattern of expression of HPT activity observed with IPNS in untransformed strains.
Asunto(s)
Acremonium/genética , Cinamatos , Fosfotransferasas (Aceptor de Grupo Alcohol) , Regiones Promotoras Genéticas , Transformación Genética , Southern Blotting , Resistencia a Medicamentos/genética , Electroforesis en Gel de Campo Pulsado , Glucosafosfato Deshidrogenasa/genética , Higromicina B/análogos & derivados , Higromicina B/farmacología , Oxidorreductasas/genética , Fosfotransferasas/genética , Fosfotransferasas/metabolismoRESUMEN
The effect of introducing user fees on the frequency and quality of MEDLINE searching with GRATEFUL MED by physicians in clinical settings was tested. After training and free use (prior study), consenting participants were randomly allocated to pay searching costs (pay group) or continue without fees (no pay group). Fifty-nine physicians participated. Among the prior study's frequent searchers, the pay group searched at less than one third of the rate of those assigned to no pay. For less frequent searchers in the prior study, only 48% of those assigned to pay did any searches, compared with 85% for the no pay group (P = 0.006), and for those who did search, their frequency was almost half. However, there was no significant difference in the quality of searches; both groups demonstrated about equivalent recall (P = 0.77), but significantly lower precision (P = 0.03) than for the librarian's independent searches. Similarly, there was no difference in the proportion of searches affecting clinical decisions for the two groups. Thus, imposing user charges for online searching in clinical settings after a period of free use adversely affects searching quantity, but not quality. MEDLINE providers should consider whether user fees will undermine its benefits.
Asunto(s)
MEDLINE/economía , Honorarios y Precios , Humanos , Bibliotecólogos , MEDLINE/estadística & datos numéricos , Ontario , Médicos , Distribución AleatoriaRESUMEN
Using a Latin square design, 4 patients were examined by 4 rheumatologists. Joints were scored for tenderness and inflammation. The Ritchie, the index of the American Rheumatism Association (ARA), the Hart modified Ritchie and a simplified Lansbury index were calculated from the raw data. The results suggest that an articular index consisting of a simple count of tender joints (Hart modified Ritchie) or a simple count of tender or swollen joints (ARA index) are the most reproducible with multiple observers. We suggest that these indices would be most appropriate for multicenter clinical trials.
Asunto(s)
Artritis Reumatoide/patología , Índice de Severidad de la Enfermedad , Humanos , Articulaciones/patologíaRESUMEN
A number of A. chrysogenum strains from a lineage improved in cephalosporin C production were analysed by contoured-clamped homogeneous electric field gel electrophoresis (CHEF). Although antibiotic titre was increased across the lineage, chromosome rearrangements were only observed at two points in it. In one member of the lineage the chromosomal changes included those which altered the size of the chromosome on which the isopenicillin N synthetase gene (pcbC) was located. It is proposed that chromosome changes are a chance event in an industrial strain improvement programme.
Asunto(s)
Acremonium/genética , Cefalosporinas/biosíntesis , Cromosomas Fúngicos/fisiología , Reordenamiento Génico , Acremonium/enzimología , Acremonium/metabolismo , Southern Blotting , ADN de Hongos/genética , ADN de Hongos/aislamiento & purificación , Genes Fúngicos , Hibridación de Ácido Nucleico , Oxidorreductasas/genéticaRESUMEN
We compared the benefits and risks of combinations of long acting antirheumatic drugs with those of the same drugs used singly in the treatment of rheumatoid arthritis. We searched the literature through MEDLINE (1966-89), Index Medicus (1956-65), Excerpta Medica (1982-89), Science Citation Index (1982-89), and bibliographic review of located articles. Of a total of 341 citations, we identified 7 prospective trials that specifically addressed the stated purpose. We independently assessed the quality of the selected trials, using published methodological criteria and summarized the effect of treatment on arthritis activity and the incidence of side effects. The trials we evaluated tested various drug combinations. Because of deficiencies in methods and reporting, only 3 trials had sufficient quality to yield strong or moderately strong evidence. None conclusively demonstrated benefit of a drug combination: 2 suggested such benefit, including 1 also suggesting increased toxicity; the 3rd suggested only increased toxicity. The other 4 trials yielded weak evidence to support both increased efficacy and toxicity. The advantages of any antirheumatic drug combination remain unproven. Because these advantages are likely to be modest, they can only be shown in rigorously designed trials enrolling large numbers of patients. Methods and reporting of antirheumatic drug trials should be standardized to allow combining of study results.
Asunto(s)
Antiinflamatorios/uso terapéutico , Artritis Reumatoide/tratamiento farmacológico , Artritis Reumatoide/epidemiología , Ensayos Clínicos como Asunto , Estudios de Cohortes , Preparaciones de Acción Retardada , Quimioterapia Combinada , Humanos , Estudios Prospectivos , Ensayos Clínicos Controlados Aleatorios como AsuntoRESUMEN
OBJECTIVE: To assist with educational planning we surveyed health sciences faculty members in 1989 to determine their use of microcomputers, desire for further instruction and perceptions on what microcomputer services should be provided for students. The 1989 results were compared with those of a similar survey performed in 1986. DESIGN: A self-completed, mailed questionnaire, with up to three reminders. SETTING: Faculty of Health Sciences, McMaster University, Hamilton, Ont. PARTICIPANTS: All full-time (FT) and part-time (PT) faculty members were sent the questionnaire; over 80% of the FT and 65% of the PT faculty members responded in 1986 and in 1989. RESULTS: The proportions of faculty members who used microcomputers increased significantly over the 3 years, from 71% to 87% among FT members (p = 2.2 x 10(-8)) and from 48% to 69% among PT members (p = 4.9 x 10(-8)). There were significant increases in the use of many of the applications, especially database and filing uses (from 10% to 41% among FT members [p less than 1 x 10(-9)] and from 6% to 34% among PT members [p less than 1 x 10(-9)]) and on-line access to bibliographic databases (from 7% to 37% among FT members [p less than 1 x 10(-9)] and from 3% to 18% among PT members [p less than 1 x 10(-9)]. These changes occurred mainly through individual initiative and voluntary continuing education. CONCLUSIONS: The extraordinary rate of adoption of microcomputers attests to their perceived usefulness. Curriculum planners need to consider how the success of microcomputer applications can be evaluated objectively and how successful applications can be integrated into educational programs.
Asunto(s)
Actitud del Personal de Salud , Actitud hacia los Computadores , Docentes , Empleos en Salud/educación , Microcomputadores/estadística & datos numéricos , Curriculum , Humanos , Ontario , Encuestas y CuestionariosRESUMEN
The objective of this study was to determine the quality of MEDLINE searches done by physicians, physician trainees, and expert searchers (clinicians and librarians). Its design was an analytic survey with independent replication in a setting of self-service online searching from medical wards, an intensive care unit, a coronary care unit, an emergency room, and an ambulatory clinic in a 300-bed teaching hospital. Participating were all M.D. clinical clerks, house, and attending staff responsible for patients in the above settings. Intervention for all participants consisted of a 2-h small group class and 1-h practice session on MEDLINE searching (GRATEFUL MED) before free access to MEDLINE. Search questions from 104 randomly selected novice searches were given to 1 of 13 clinicians with prior search experience and 1 of 3 librarians to run independent searches (triplicated searches). Measurements and main results from these unique citations of the triplicated searches were sent to expert clinicians to rate for relevance (7-point scale). Recall (number of relevant citations retrieved from an individual search divided by the total number of relevant citations from all searches on the same topic) and precision (proportion of relevant citations retrieved in each search) were calculated. Librarians were significantly better than novices for both. Librarians had equivalent recall to, and better precision than, experienced end-users. Unexpectedly, only 20% of relevant citations were retrieved by more than one search of the set of three, with the conclusion that novice searchers on MEDLINE via GRATEFUL MED after brief training have relatively low recall and precision. Recall improves with experience but precision remains suboptimal. Further research is needed to determine the "learning curve," evaluate training interventions, and explore the non-overlapping retrieval of relevant citations by different searchers.
Asunto(s)
MEDLINE , Estudios de Evaluación como Asunto , Grateful Med , Reproducibilidad de los Resultados , Descriptores , Interfaz Usuario-ComputadorRESUMEN
We report the development of a homologous transformation system for Cephalosporium acremonium using the niaD gene of the nitrate assimilation (NA) pathway. Mutants in the NA pathway were selected on the basis of chlorate resistance by conventional means. Screening procedures were developed to differentiate between nitrate reductase apoprotein structural gene mutants (niaD) and molybdenum cofactor gene mutants (cnx) as wt C. acremonium, unlike most filamentous fungi, fails to grow on minimal medium with hypoxanthine as a sole source of nitrogen. Phage clones carrying the niaD gene were isolated from a C. acremonium library constructed in lambda EMBL3 using the A. nidulans niaD gene as a heterologous probe. An 8.6-kb EcoRI fragment was subcloned into pUC18, and designated pSTA700. pSTA700 was able to transform stable niaD mutants to NA at a frequency of up to 40 transformants per microgram DNA. Transformants were easily visible since the background growth was low and no abortives were observed. Gene replacements, single copy homologous integration and complex multiple integrations were observed. The niaD system was used to introduce unselected markers for hygromycin B resistance and benomyl resistance into C. acremonium by cotransformation.
Asunto(s)
Acremonium/genética , Coenzimas/genética , Genes Fúngicos , Metaloproteínas/metabolismo , Molibdeno/metabolismo , Nitrato Reductasas/genética , Pteridinas/metabolismo , Transformación Genética , Acremonium/efectos de los fármacos , Acremonium/enzimología , Acremonium/crecimiento & desarrollo , Benomilo/farmacología , Cloratos/farmacología , Farmacorresistencia Microbiana , Frecuencia de los Genes , Marcadores Genéticos , Higromicina B/farmacología , Cofactores de Molibdeno , Mutación , Nitrato-Reductasa , Nitrato Reductasas/biosíntesis , Mapeo RestrictivoRESUMEN
Total RNA was extracted daily from the beta-lactam antibiotic producing fungus A. chrysogenum strain CO728 during a 7 day cephalosporin C fermentation. IPNS mRNA species, with a size of about 1.5 kb, were detected by Northern blotting at high levels between days 2 and 4. The rapid appearance of IPNS mRNA in mycelial extracts up to day 2 suggests that IPNS is regulated at the transcriptional level. Primer extension and S1 endonuclease mapping studies indicate the existence of two major and at least two minor transcription initiation start sites. There was no change in the relative levels of the four transcripts during the period they could be detected. A region upstream of the IPNS structural gene (pcbC) has been sequenced and the transcription initiation sites appear as major and minor pairs on either side of one of the pyrimidine-rich blocks that punctuate the promoter sequence.