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71.
Wayne W. Grody Deborah Klein Amy E. Dodson Rita M. Kern Paul B. Wissmann Barbara K. Goodman Patrick Bassand Bert Marescau Soo-Sang Kang James V. Leonard Stephen D. Cederbaum 《American journal of human genetics》1992,50(6):1281-1290
We have explored the molecular pathology in 28 individuals homozygous or heterozygous for liver arginase deficiency (hyperargininemia) by a combination of Southern analysis, western blotting, DNA sequencing, and PCR. This cohort represents the majority of arginase-deficient individuals worldwide. Only 2 of 15 homozygous patients on whom red blood cells were available had antigenically cross-reacting material as ascertained by western blot analysis using anti-liver arginase antibody. Southern blots of patient genomic DNAs, cut with a variety of restriction enzymes and probed with a near-full-length (1,450-bp) human liver arginase cDNA clone, detected no gross gene deletions. Loss of a TaqI cleavage site was identified in three individuals: in a homozygous state in a Saudi Arabian patient at one site, at a different site in homozygosity in a German patient, and in heterozygosity in a patient from Australia. The changes in the latter two were localized to exon 8, through amplification of this region by PCR and electrophoretic analysis of the amplified fragment after treatment with TaqI; the precise base changes (Arg291X and Thr290Ser) were confirmed by sequencing. It is interesting that the latter nucleotide variant (Thr290Ser) was found to lie adjacent to the TaqI site rather than within it, though whether such a conservative amino acid substitution represents a true pathologic mutation remains to be determined. We conclude that arginase deficiency, though rare, is a heterogeneous disorder at the genotypic level, generally encompassing a variety of point mutations rather than substantial structural gene deletions. 相似文献
72.
Raffaella Scardigli Silvia Soddu Rita Falcioni Marco Crescenzi Letizia Cimino Ada Sacchi 《Experimental cell research》1996,227(2):223
Integrin β4subunit is present in association with α6chain on both normal and transformed epithelial cells. Recently α6β4heterodimer was found on the endothelium of medium-sized blood vessels and on immature thymocytes. In this report we show, by Northern blotting, indirect immunofluorescence, immunoprecipitation, and Western blotting, that β4subunit is expressed also on cells of mesenchymal origin such as fibroblasts, myoblasts, and myotubes. Increased expression of α6β4has been related to the aggressive metastatic phenotype of human and murine carcinomas. The transforming growth factor β1(TGF-β1) has been found to modulate the expression of several integrins and intracellular matrix proteins, as well as to stimulate cell invasion and metastatic potential. To evaluate whether α6β4expression is modulated by TGF-β1, we transfected 3T3 fibroblasts with an expression vector carrying the human TGF-β1cDNA driven by the SV40 early promoter. We observed by indirect immunofluorescence a modification in the subcellular distribution of β4subunit, which acquires a perinuclear localization. This finding suggests this integrin subunit correlates with the cytoskeletal reorganization induced by TGF-β1. 相似文献
73.
L. Ferraro S. Tanganelli L. Marani C. Bianchi L. Beani A. Siniscalchi 《Neurochemical research》1996,21(5):547-552
The effects of α-glycerylphosphorylcholine (α-GPC) on endogenous cortical GABA release were studied both in vivo and in vitro.
In freely moving rats, equipped with epidural cups, α-GPC (30–300 mg/kg i.p.) increased GABA release. This effect was potentiated
by atropine, both systematically administered (5 mg/kg i.p.) and locally applied (1.4 μM), but not by mecamylamine (4 mg/kg
i.p.). The α-GPC-induced increasein GABA release was abolished in rats pretreated with the α1 receptor antagonist prazosin (14 μg/kg i.p.). In cortical slices α-GPC (0.4 mM) increased the spontaneous GABA efflux. This effectwas abolished by tetrodotoxin (0.5 μM) and prazosin (1 μM), but not by atropine (0.15 μM) ormecamylamine (2.5μM). These results indicate that the facilitatory response by α-GPC on GABArelease does not depend on a direct activation of either muscarinic or nicotinic receptors, but suggest the involvement of the noradrenergic
system. 相似文献
74.
The low percentage of living bacteria commonly obtained when comparing viable counts with total direct counts in seawater could be due more to inappropriate techniques for appreciating the growth ability of living cells than to unadapted culture conditions. The most-probable-number counts in filtered seawater cultures and the microscopic counts of 4(prm1),6-diamidino-2-phenylindole (DAPI)-stained aggregate-forming units grown on black polycarbonate filters appeared significantly correlated to the direct counts. Both these techniques show that in the superficial and intermediate water masses, the living cells may constitute an important (frequently higher than 20%) but highly variable part of the total populations. These viable counts appear more realistic than the conventional CFU counts, which provide only 0.001 to 0.2% of the total counts. 相似文献
75.
The presence of bacteria in a deep clay sediment was analyzed in a 20-m-long core horizontally drilled from a mine gallery at a depth of 224 m in the Boom clay formation (Mol, Belgium). This clay deposit is the result of a marine sedimentary process that occurred 35 million years ago. Bacterial activities were estimated by measuring respiration on [14C]glucose. Using the same samples, universal primers for the genes coding for eubacterial 16S rRNA were used to amplify extracted DNA. PCR products were then cloned, sequenced, and analyzed by molecular phylogeny. Our data showed a decrease in bacterial densities as a function of distance from the gallery, with few bacteria detectable by culture at more than 80 cm from the gallery wall. PCR experiments showed the presence of bacteria in all samples, and phylogenetic analyses were then used to tentatively identify these organisms. Because of low bacterial densities in deep clay samples, direct counts and enumeration of viable bacteria on diverse culture media remained negative. All experiments, both cultures and PCR, demonstrated the difficulty of analyzing samples that contain only a few poorly active bacteria as it is difficult to avoid a small contamination by active bacteria during sampling. Since the porosity of the Boom clay formation is less than the expected size of bacteria, it is possible that some of the bacteria present in this 35-million-year-old deep clay deposit derive from cells initially trapped during the sedimentation process. 相似文献
76.
G. Bestetti P. Di Gennaro E. Galli B. Leoni F. Pelizzoni G. Sello D. Bianchi 《Applied microbiology and biotechnology》1994,40(6):791-793
The Pseudomonas fluorescens N3 was isolated from soil for its ability to utilize naphthalene as a carbon source. The strain transforms 2,3-dimethyl-, 2-methoxy-, 1- and 2-ethylnaphthalenes to the corresponding salicylic acids competitively with chemical synthesis. The identification of 2-hydroxy-2-carboxy-7-ethylchromane by biotransformation of 2-ethylnaphthalene, contributes to elucidating the steps involved in the catabolic pathways of naphthalenes to salicylaldehydes.
Correspondence to: F. Pelizzoni 相似文献
77.
Multiple DNA Variant Association Analysis: Application to the Insulin Gene Region in Type 1 Diabetes 下载免费PDF全文
C. Julier A. Lucassen P. Villedieu M. Delepine C. Levy-Marchal P. M. Danz F. Bianchi C. Boitard P. Froguel J. Bell G. M. Lathrop 《American journal of human genetics》1994,55(6):1247-1254
Association and linkage studies have shown that at least one of the genetic factors involved in susceptibility to insulin-dependent diabetes mellitus (IDDM) is contained within a 4.1-kb region of the insulin gene. Sequence analysis has led to the identification of 10 DNA variants in this region that are associated with increased risk for IDDM. These variants are in strong linkage disequilibrium with each other, and previous studies have failed to distinguish between the variant(s) that cause increased susceptibility to IDDM and others that are associated with the disease because of linkage disequilibrium. To address this problem, we have undertaken a large population study of French diabetics and controls and have analyzed genotype patterns for several of the variant sites simultaneously. This has led to the identification of a subset consisting of four variants (−2733AC, −23HphI, −365VNTR, and +1140AC), at least one of which appears to be directly implicated in disease susceptibility. The multiple-DNA-variant association-analysis approach that is applied here to the problem of identifying potential susceptibility variants in IDDM is likely to be important in studies of many other multifactorial diseases. 相似文献
78.
Ichiro Nakayama Fausto Foresti Rita Tewari Manfred Schartl Daniel Chourrout 《Chromosoma》1994,103(1):31-39
In order to study the divergence of teleost sex chromosomes, subtractive cloning was carried out between genomic DNA of males and females of the rainbow trout (XX/XY) and of Leporinus elongatus (ZW/ZZ). Inserts cloned in a plasmid vector were individually tested on Southern blots of DNA of males and females for sex specificity. No sex-specific insert was obtained from trout, but two out of ten inserts cloned from L. elongatus showed sex-specific patterns in this species: one corresponds to a sequence present on both Z and W chromosomes, while the other is W specific. Sequences of these two inserts show neither clear homology with other known sequences, nor an open reading frame. They cross-hybridize with the genomic DNA of Leporinus friderici, but without sex-specific patterns. Twenty-four L. elongatus adults were sexed by gonadal observation, chromosomed examination and Southern hybridization with one or the other insert. Ten males and 11 females had chromosomes and hybridization patterns typical of their sex. One ZW female was recognized as a male with the W-specific probe. This was also the case for two unusual ZW males, one having a male hybridization pattern with the other probe. These three atypical individuals may result from single genetic exchanges between four regions of the Z and the W, giving rise to three atypical W chromosomes. Finding males with such atypical heterochromosomes in a female heterogametic species may indicate that a gradual transition occurs between the heterogametic systems. 相似文献
79.
Maria Rita Santi Stefano Vicini †Basil Eldadah Joseph H. Neale 《Journal of neurochemistry》1994,63(6):2357-2360
Abstract: With the use of the single-cell polymerase chain reaction (PCR), the GABAA receptor subunit mRNA content was analyzed in granule and Purkinje neurons from rat cerebellar slices. We used an experimental protocol to assess simultaneously the presence of two subunits in each cell while electrophysiological recordings were performed with the whole-cell patch-clamp technique. Based on a computer alignment of the nucleotide sequence corresponding to α1 and α6 GABAA receptor subunits, homologous regions were identified that allowed coamplification of both mRNAs using a single primer combination. The presence of selective restriction sites within the targeted templates allowed us to identify which receptor subunit mRNAs were coamplified by performing restriction enzyme-mediated cleavage of the amplification products. In all Purkinje neurons assayed, α1 subunit mRNA but not α6 mRNA was detected. In contrast, among individual granule neurons we found a heterogeneous distribution of the mRNA for the α1 and α6 GABAA receptor subunits. A comparison of the results of the PCR amplification and the analysis of GABA-mediated inhibitory synaptic currents does not allow us to identify kinetic characteristics of synaptic currents that clearly correlate with the presence or the absence of α6 subunit mRNA. 相似文献
80.