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31.
P. Portegies J. de Gans J. M. Lange M. M. Derix H. Speelman M. Bakker S. A. Danner J. Goudsmit 《BMJ (Clinical research ed.)》1989,299(6703):819-821
OBJECTIVE--To assess the incidence of the AIDS dementia complex and the presence of HIV I p24 antigen in cerebrospinal fluid in relation to zidovudine treatment. DESIGN--Retrospective study of a consecutive series of patients with AIDS from 1982 to 1988. SETTING--An academic centre for AIDS. PATIENTS--196 Patients with AIDS and neurological symptoms examined from 1982 to 1988. INTERVENTIONS--Zidovudine treatment, which was introduced to The Netherlands on 1 May 1987 for patients with severe symptoms of HIV infection (Centers for Disease Control groups IVA, B, C, and D). MAIN OUTCOME MEASURES--Diagnosis of AIDS dementia complex and presence of HIV I p24 antigen in cerebrospinal fluid. RESULTS--The AIDS dementia complex was diagnosed in 40 of the 196 (20%) patients with AIDS. Thirty eight of 107 patients with AIDS (36%) not taking zidovudine developed the AIDS dementia complex compared with two of the 89 (2%) taking the drug (p less than 0.00001). The incidence of the AIDS dementia complex increased to 53% in the first half of 1987, after the introduction of zidovudine in May 1987, decreasing to 10% in the second half of 1987 and to 3% in 1988. Dementia was diagnosed before definition of the AIDS dementia complex (1986) according to DSM-III criteria and there was good agreement between diagnosis before and after 1986. Sixteen of 61 samples of cerebrospinal fluid (26%) from patients with AIDS (10 with the AIDS dementia complex) not taking zidovudine were positive for HIV I p24 antigen, whereas none of 37 cerebrospinal fluid samples from patients with AIDS (two with the AIDS dementia complex) taking zidovudine were positive. CONCLUSIONS--The incidence of AIDS dementia complex in patients with AIDS declined after the introduction of systematic treatment with zidovudine; the AIDS dementia complex might be prevented by inhibiting viral replication in the central nervous system. 相似文献
32.
High resolution deletion breakpoint mapping in the DMD gene by whole cosmid hybridization. 总被引:16,自引:1,他引:15
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L A Blonden J T den Dunnen H M van Paassen M C Wapenaar P M Grootscholten H B Ginjaar E Bakker P L Pearson G J van Ommen 《Nucleic acids research》1989,17(14):5611-5621
The locus DXS269 (P20) defines a deletion hotspot in the distal part of the Duchenne Muscular Dystrophy gene. We have cloned over 90 kilobase-pairs of genomic DNA from this region in overlapping cosmids. The use of whole cosmids as probes in a competitive DNA hybridization analysis proves a fast and convenient method for identifying rearrangements in this region. A rapid survey of P20-deletion patients is carried out to elucidate the nature of the propensity to deletions in this region. Using this technique, deletion breakpoints are pinpointed to individual restriction fragments in patient DNAs without the need for tedious isolation of single copy sequences. Simultaneously, the deletion data yield a consistent restriction map of the region and permit detection of several RFLPs. A 176 bp exon was identified within the cloned DNA, located 3' of an intron exceeding 150 Kb in length. Its deletion causes a frameshift in the dystrophin reading frame and produces the DMD phenotype. This exon is one of the most frequently deleted exons in BMD/DMD patients and its sequence is applied in a pilot study for diagnostic deletion screening using Polymerase Chain Reaction amplification. 相似文献
33.
Detection and identification of FaeC as a minor component of K88 fibriilae of Escherichia coli 总被引:1,自引:0,他引:1
B. Oudega M. de Graaf L. de Boer D. Bakker C. E. M. Vader F. R. Mooi F. K. de Graaf 《Molecular microbiology》1989,3(5):645-652
A tribrid gene containing ompF, faeC, and lacZ sequences was constructed by subcloning a large central segment of the K88ab gene encoding the fibrillar subunit-like protein FaeC into the open reading frame expression vector pORF2. The resulting tribrid protein was isolated and used to raise antibodies against the FaeC protein. These antibodies were then used for the detection and subcellular localization of the FaeC protein in Escherichia coli harbouring the K88ab-encoding plasmid pFM205 or mutant derivatives. Immunoblotting of subcellular fractions and of purified fibrillae, and agglutination experiments using whole cells revealed that the FaeC protein is present in the periplasm and as a minor component in the K88ab fibrillae. FaeC was also detected in purified K88ac and K88ad fibrillae. Immunoelectron microscopy confirmed the presence of FaeC in K88ab fibrillae, particularly at the tips of the longer fibrillae. 相似文献
34.
Methylation of GATC sites is required for precise timing between rounds of DNA replication in Escherichia coli. 总被引:16,自引:7,他引:9
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We have used the Koppes and Nordstr?m (Cell 44:117-124, 1986) CsCl density transfer approach for analysis of DNA from exponentially growing, isogenic Escherichia coli dam+ and dam mutant cells to show that timing between DNA replication initiation events is precise in the dam+ cells but is essentially random in the dam cells. Thus, methylation of one or more GATC sites, such as those found in unusual abundance within the origin, oriC, is required for precise timing between rounds of DNA replication, and precise timing between initiation events is not required for cell viability. Both the dam-3 point mutant and the delta(dam)100 complete deletion mutant were examined. The results were independent of the mismatch repair system; E. coli mutH cells showed precise timing, whereas timing in the isogenic E. coli mutH delta(dam)100 double mutant was random. The mechanism is thus different from the role of Dam methylation in mismatch repair and probably involves conversion of hemimethylated GATC sites present in daughter origins just after initiation to a fully methylated state. 相似文献
35.
Different mosaicism frequencies for proximal and distal Duchenne muscular dystrophy (DMD) mutations indicate difference in etiology and recurrence risk. 总被引:8,自引:0,他引:8
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M R Passos-Bueno E Bakker A L Kneppers R I Takata D Rapaport J T den Dunnen M Zatz G J van Ommen 《American journal of human genetics》1992,51(5):1150-1155
In about 65% of the cases of Duchenne muscular dystrophy (DMD) a partial gene deletion or duplication in the dystrophin gene can be detected. These mutations are clustered at two hot spots: 30% at the hot spot in the proximal part of the gene and about 70% at a more distal hot spot. Unexpectedly we observed a higher frequency of proximal gene rearrangements among proved "germ line" mosaic cases. Of the 24 mosaic cases we are aware of, 19 (79%) have a proximal mutation, while only 5 (21%) have a distal mutation. This finding indicates that the mutations at the two hot spots in the dystrophin gene differ in origin. Independent support for the different mosaicism frequency was found by comparing the mutation spectra observed in isolated cases of DMD and familial cases of DMD. In a large two-center study of 473 patients from Brazil and the Netherlands, we detected a significant difference in the deletion distribution of isolated (proximal:distal ratio 1:3) and familial cases (ratio 1:1). We conclude from these data that proximal deletions most likely occur early in embryonic development, causing them to have a higher chance of becoming familial, while distal deletions occur later and have a higher chance of causing only isolated cases. Finally, our findings have important consequences for the calculation of recurrence-risk estimates according to the site of the deletion: a "proximal" new mutant has an increased recurrence risk of approximately 30%, and a "distal" new mutant has a decreased recurrence risk of approximately 4%. 相似文献
36.
Significant linkage disequilibrium between the Huntington disease gene and the loci D4S10 and D4S95 in the Dutch population.
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M I Skraastad E Van de Vosse R Belfroid K Hld M Vegter-van der Vlis L A Sandkuijl E Bakker G J van Ommen 《American journal of human genetics》1992,51(4):730-735
Significant linkage disequilibrium has been found between the Huntington disease (HD) gene and DNA markers located around D4S95 and D4S98. The linkage-disequilibrium studies favor the proximal location of the HD gene, in contrast to the conflicting results of recombination analyses. We have analyzed 45 Dutch HD families with 19 DNA markers and have calculated the strength of linkage disequilibrium. Highly significant linkage disequilibrium has been detected with D4S95, consistent with the studies in other populations. In contrast with most other studies, however, the area of linkage disequilibrium extends from D4S10 proximally to D4S95, covering 1,100 kb. These results confirm that the HD gene most likely maps near D4S95. 相似文献
37.
Effect of pseudobactin 358 production by Pseudomonas putida WCS358 on suppression of fusarium wilt of carnations by nonpathogenic Fusarium oxysporum Fo47. 总被引:2,自引:2,他引:0
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P Lemanceau P A Bakker W J De Kogel C Alabouvette B Schippers 《Applied microbiology》1992,58(9):2978-2982
Nonpathogenic Fusarium oxysporum Fo47b10 combined with Pseudomonas putida WCS358 efficiently suppressed fusarium wilt of carnations grown in soilless culture. This suppression was significantly higher than that obtained by inoculation of either antagonistic microorganism alone. The increased suppression obtained by Fo47b10 combined with WCS358 only occurred when Fo47b10 was introduced at a density high enough (at least 10 times higher than that of the pathogen) to be efficient on its own. P. putida WCS358 had no effect on disease severity when inoculated on its own but significantly improved the control achieved with nonpathogenic F. oxysporum Fo47b10. In contrast, a siderophore-negative mutant of WCS358 had no effect on disease severity even in the presence of Fo47b10. Since the densities of both bacterial strains at the root level were similar, the difference between the wild-type WCS358 and the siderophore-negative mutant with regard to the control of fusarium wilt was related to the production of pseudobactin 358. The production of pseudobactin 358 appeared to be responsible for the increased suppression by Fo47b10 combined with WCS358 relative to that with Fo47b10 alone. 相似文献
38.
A new principle of cell sorting by using selective electroporation in a modified flow cytometer 总被引:2,自引:0,他引:2
When a strong electric field pulse of a few microseconds is applied to biological cells, small pores are formed in the cell membranes; this process is called electroporation. At high field strengths and/or long pulse durations the membranes will be damaged permanently. This eventually leads to cell kill. We have developed a modified flow cytometer in which one can electroporate individual cells selected by optical analysis. The first experiments with this flow cytometer were designed to use it as a damaging sorter; we used electric pulses of 10 microseconds and resulting field strengths of 2.0 and 3.2 x 10(6) V/m to kill K562 cells and lymphocytes respectively. The hydrodynamically focused cells are first optically analyzed in the usual way in a square flow channel. At the end of this channel the cells are forced to flow through a small Coulter orifice, into a wider region. If optical analysis indicates that a cell is unwanted, the cell is killed by applying a strong electric field across the Coulter orifice. The wanted living cells can be subsequently separated from the dead cells and cell fragments by a method suitable for the particular application (e.g., centrifugation, cell growth, density gradient, etc.). The results of these first experiments demonstrate that by using very simple equipment, sorting by selective killing with electric fields is possible at rates of 1,000 cells/s with a purity of the sorted fraction of 99.9%. 相似文献
39.
Alexander Schouten Jan Roosien Fred A. van Engelen G. A. M. de Jong A. W. M. Borst-Vrenssen Jacoline F. Zilverentant Dirk Bosch Willem J. Stiekema Fred J. Gommers Arjen Schots Jaap Bakker 《Plant molecular biology》1996,30(4):781-793
The effects of subcellular localization on single-chain antibody (scFv) expression levels in transgenic tobacco was evaluated using an scFv construct of a model antibody possessing different targeting signals. For translocation into the secretory pathway a secretory signal sequence preceded the scFv gene (scFv-S). For cytosolic expression the scFv antibody gene lacked such a signal sequence (scFv-C). Also, both constructs were provided with the endoplasmic reticulum (ER) retention signal KDEL (scFv-SK and scFv-CK, respectively). The expression of the different scFv constructs in transgenic tobacco plants was controlled by a CaMV 35S promoter with double enhancer. The scFv-S and scFv-SK antibody genes reached expression levels of 0.01% and 1% of the total soluble protein, respectively. Surprisingly, scFv-CK transformants showed considerable expression of up to 0.2% whereas scFv-C transformants did not show any accumulation of the scFv antibody. The differences in protein expression levels could not be explained by the steady-state levels of the mRNAs. Transient expression assays with leaf protoplasts confirmed these expression levels observed in transgenic plants, although the expression level of the scFv-S construct was higher. Furthermore, these assays showed that both the secretory signal and the ER retention signal were recognized in the plant cells. The scFv-CK protein was located intracellularly, presumably in the cytosol. The increase in scFv protein stability in the presence of the KDEL retention signal is discussed. 相似文献
40.
Hybridomas were cultured under steady-state conditions in a series of two continuous stirred-tank reactors (CSTRs), using a serum-free medium. The substrate not completely converted in the first CSTR, was transported with the cells to the second one and very low growth rates, high death rates, and lysis of viable cells were observed in this second CSTR. These conditions are hardly accessible in a single vessel, because such experiments would be extremely time-consuming and unstable due to a low viability. In contrast to what is often observed in literature, kinetic parameters could thus be derived without the neccessity for extrapolation to lower growth rates. Good agreement with literature averages for other hybridomas was found. Furthermore, showing that the reactor series is a valuable research tool for kinetic studies under extreme conditions, the possibility to observe cell death under stable and defined steady-state conditions offers interesting opportunities to investigate apoptosis and necrosis. Additionally, a model was developed that describes hybridoma growth and monoclonal antibody production in the bioreactor cascade on the basis of glutamine metabolism. Good agreement between the model and the experiments was found.Abbreviation MAb
Monoclonal antibody
Nomenclature
C
AConcentration of any (mol m-3) component A
D Dilution rate (s-1)
K
dDeath-rate constant (mol m-3)
K
lLysis-rate constant (mol m-3)
K
sMonod constant (mol m-3)
m Maintenance coefficient (mol cell-1 s-1)
q Specific consumption (mol cell-1 s-1) or production rate
t Time (s)
X Cell concentration (cell m-3)
Y Yield coefficient (cell mol-1)
Greek symbols
d
Specific death rate (s-1)
l
Specific lysis rate (s-1) of viable cells
net
Net specific growth (s-1) rate
true
True specific growth (s-1) rate 相似文献