Polyacrylamide–agarose gel electrophoretic analysis of tobacco mosaic virus disassembly intermediates

1984 ◽  
Vol 62 (11) ◽  
pp. 2336-2339 ◽  
Author(s):  
Richard Hogue ◽  
Alain Asselin
Keyword(s):  
Tobacco Mosaic Virus ◽  
Mosaic Virus ◽  
Gel Electrophoresis ◽  
Electrophoretic Analysis ◽  
Sucrose Density Gradient ◽  
Plant Viruses ◽  
Buffer System ◽  
Agarose Gel ◽  
Agarose Gel Electrophoresis

The intermediates of disassembly of the U1, U4, U6, and U7 strains of tobacco mosaic virus (TMV) induced by alkali and urea were directly analysed by electrophoresis in composite polyacrylamide (2.0 to 2.5% (w/v)) – agarose (0.5% (w/v)) gels, using a discontinuous buffer system with Tris, borate, EDTA, and sodium chloride (pH 8.3). The results show no difference between the disassembly patterns of the four strains. In the case of alkaline stripping, two new major partially stripped virus (PSV) particles (PSV 5a and PSV 5c) and several minor intermediates were characterized. The overall results indicate that polyacrylamide–agarose gel electrophoresis is superior to agarose gel electrophoresis and to sucrose density gradient ccntrifugation for the analytical separation of PSVs. Polyacrylamide–agarose gel electrophoresis is thus a rather simple, rapid, and efficient method of studying in vitro disassembly of plant viruses.

Study of tobacco mosaic virus in vitro disassembly by sucrose density gradient centrifugation and agarose gel electrophoresis

1984 ◽  
Vol 62 (3) ◽  
pp. 457-462 ◽  
Author(s):  
Richard Hogue ◽  
Alain Asselin
Keyword(s):  
Tobacco Mosaic Virus ◽  
Mosaic Virus ◽  
Density Gradient ◽  
Gel Electrophoresis ◽  
Density Gradient Centrifugation ◽  
Gradient Centrifugation ◽  
Agarose Gel ◽  
Agarose Gel Electrophoresis ◽  
Sucrose Density Gradient Centrifugation

In vitro disassembly of tobacco mosaic virus (TMV) strains U1, U2, U4, U6, and U7 with alkali and urea was studied by sucrose or sucrose–dimethylsulfoxide (DMSO) density gradient centrifugation and by agarose gel electrophoresis. All strains gave similar decapsidation patterns with both agents when partially stripped virus particles (PSVs) were analyzed by sedimentation and electrophoresis. However, U6 was more sensitive to decapsidation than the other strains and U2 exhibited resistance to decapsidation. Agarose gel electrophoresis of TMV decapsidation products allowed the detection of several classes of PSVs in addition to aggregation products involving PSVs and monomer particles. Agarose gel electrophoresis is thus very rapid and useful for analysis of TMV disassembly products especially when aggregation phenomena and kinetic studies with numerous samples are considered.

Characterisation Of Active Components Of Fix Concentrates Used In FVIII Inhibitor Patients

1981 ◽  
Author(s):  
M J Seghatchian ◽  
I J Mackie
Keyword(s):  
Gel Electrophoresis ◽  
Molecular Form ◽  
Agarose Gel ◽  
Agarose Gel Electrophoresis ◽  
Active Components ◽  
Fviii Inhibitor ◽  
Antigenic Sites ◽  
Twin Peak

It is well recognised that haemostasis may be achieved in patients with FVIII inhibitors using FIX concentrates (PTX). However, it has not been possible to correlate the clinical response with enzymicity, FVIII levels and other in vitro results. This is partially due to the presence of variable amounts of inhibitors and stabilizers such as: citrate, heparin, and anti proteases in different preparations. Agarose gel electrophoresis was performed in an attempt to separate these inhibitors so that the various activities may be measured relative to one another. PTX from 4 sources were used in this study,(Kabi Vitrum, Immuno, Oxford and Edinburgh fractionation centres); FVIII was measured by two stage clotting(FVIII:C)and amidolytic (CAM) assays, and its electrophoretic distribution was followed by preincubation with 125I-anti VIII:C (VIII:CAg). Enzymicity was measured with various substrates (S2160, S2222, S2302, S2238, S2288, S2251). All concentrates showed a fast moving peak of FVIII:C and VIII:CAm activity, in amounts in excess of that detected before electrophoresis. This peak correlated well with the distribution of bound radiolabel (VIII:CAg). Preincubation of PTX with FVIII concentrate caused the VIII:CAg peak to migrate more slowly, in its normal α 2 position. FVIII:RAg was present in much lower amounts than FVIII:C. When a twin- peak of VIII:CAm activity was sometimes seen, the slower peak was associated with S2288 activity, which may represent the action of activated factors VII or IX: other substrates showed variable patterns of enzymicity but the strongest activity was with S2160 and S2302 from the well to the β region, and this could be completely inhibited by addition of anti Kallikrein. The presence of an abnormal molecular form of FVIII, and the discrepancy between pre and post electrophoresis FVIII levels, indicate that FVIII could play a major role in the action of PTX. This may involve 1) a complex with phospholipid, FIX and FX making it potentially more effective; 2) enhancement of VIII:C activity in vivo by binding to FVIII:RAg; 3) altered antigenic sites on the FVIII, making it unrecognisable to inhibitor.

scholarly journals Serum monoclonal immunoglobulin bands in undifferentiated lymphomas of Burkitt and non-Burkitt types

Blood ◽  
1983 ◽  
Vol 61 (4) ◽  
pp. 726-731
Author(s):  
I Magrath ◽  
D Benjamin ◽  
N Papadopoulos
Keyword(s):  
High Resolution ◽  
Tumor Markers ◽  
Light Chain ◽  
Gel Electrophoresis ◽  
Lymphoblastic Lymphoma ◽  
Agarose Gel ◽  
Agarose Gel Electrophoresis ◽  
Monoclonal Immunoglobulin ◽  
Potential Value

Using an improved electroimmunofixation technique that combines the sensitivity of high resolution agarose gel electrophoresis with the specificity of immunoprecipitation, we have demonstrated monoclonal immunoglobulin bands in the serum of patients with undifferentiated lymphomas of Burkitt and non-Burkitt types. Monoclonal bands were detected in the serum of 12 of 21 patients with extensive tumor, and 1 of 10 patients with minimal tumor. All of the bands were identified as IgM of a single light chain class. Such bands were not detected in the serum of patients with lymphoblastic lymphoma (7) or African Burkitt's lymphoma (6). There was disappearance of the bands after therapy and reappearance at relapse. These findings, coupled with previously reported in vitro information, indicate that undifferentiated lymphoma cells secrete immunoglobulin of IgM isotype. Therefore, such monoclonal bands may be of potential value as tumor markers.

scholarly journals Serum monoclonal immunoglobulin bands in undifferentiated lymphomas of Burkitt and non-Burkitt types

Blood ◽  
1983 ◽  
Vol 61 (4) ◽  
pp. 726-731 ◽  
Author(s):  
I Magrath ◽  
D Benjamin ◽  
N Papadopoulos
Keyword(s):  
High Resolution ◽  
Tumor Markers ◽  
Light Chain ◽  
Gel Electrophoresis ◽  
Lymphoblastic Lymphoma ◽  
Agarose Gel ◽  
Agarose Gel Electrophoresis ◽  
Monoclonal Immunoglobulin ◽  
Potential Value

Abstract Using an improved electroimmunofixation technique that combines the sensitivity of high resolution agarose gel electrophoresis with the specificity of immunoprecipitation, we have demonstrated monoclonal immunoglobulin bands in the serum of patients with undifferentiated lymphomas of Burkitt and non-Burkitt types. Monoclonal bands were detected in the serum of 12 of 21 patients with extensive tumor, and 1 of 10 patients with minimal tumor. All of the bands were identified as IgM of a single light chain class. Such bands were not detected in the serum of patients with lymphoblastic lymphoma (7) or African Burkitt's lymphoma (6). There was disappearance of the bands after therapy and reappearance at relapse. These findings, coupled with previously reported in vitro information, indicate that undifferentiated lymphoma cells secrete immunoglobulin of IgM isotype. Therefore, such monoclonal bands may be of potential value as tumor markers.

scholarly journals Curing of Escherichia coli Antibiotic Resistance Plasmids by Aspirin

2014 ◽  
Vol 8 (1) ◽  
pp. 30-35
Author(s):  
Ibtesam H. Al Musawi ◽  
Ali A. Al Zaag
Keyword(s):  
Escherichia Coli ◽  
Gel Electrophoresis ◽  
Agarose Gel ◽  
Large Plasmid ◽  
Agarose Gel Electrophoresis ◽  
E Coli ◽  
Resistance Plasmids ◽  
Minimum Number

An isolate of Escherichia coli (E. coli) was isolated from urine sample due to person infected with urinary tract infection (UTI).The isolate was resistant to the following antibiotics: Ampicillin, Cotrimoxazole, Chloramphenicol and Tetracycline. Agarose gel electrophoresis of its plasmids content has revealed the presence of single large plasmid and two small plasmids bands. The large plasmid was conjugative and contained the resistance genes for four antibiotics. In vitro curing of this plasmid was achieved by treatment with salicylic acid (aspirin) with 150,200,250 and 300 µg/ml as indicated by the elimination of resistance, also by absence of large plasmid band following agarose gel electrophoresis. In vivo curing was conducted using New Zealand rabbits. UTI was induced by bacterial inoculation via urethral catheterization. The E. coli from urine samples of the rabbits, the count of which was proportional to the type of treatment. Minimum number of colonies was associated with group treated with metheprim was aspirin 300mg/kg daily dosage. This result may indicate that the side effect of metheprim was in its maximum with aspirin. Survivor bacteria may indicate incomplete exposure to the drug.

scholarly journals The p122 Subunit of Tobacco Mosaic Virus Replicase Is a Potent Silencing Suppressor and Compromises both Small Interfering RNA- and MicroRNA-Mediated Pathways

2007 ◽  
Vol 81 (21) ◽  
pp. 11768-11780 ◽  
Author(s):  
Tibor Csorba ◽  
Aurelie Bovi ◽  
Tamás Dalmay ◽  
József Burgyán
Keyword(s):  
Tobacco Mosaic Virus ◽  
Mosaic Virus ◽  
Nuclear Export ◽  
Rna Silencing ◽  
Plant Viruses ◽  
Silencing Suppressor ◽  
Small Interfering Rnas ◽  
In Planta ◽  
Interfering Rna

ABSTRACT One of the functions of RNA silencing in plants is to defend against molecular parasites, such as viruses, retrotransposons, and transgenes. Plant viruses are inducers, as well as targets, of RNA silencing-based antiviral defense. Replication intermediates or folded viral RNAs activate RNA silencing, generating small interfering RNAs (siRNAs), which are the key players in the antiviral response. Viruses are able to counteract RNA silencing by expressing silencing-suppressor proteins. It has been shown that many of the identified silencing-suppressor proteins bind long double-stranded RNA or siRNAs and thereby prevent assembly of the silencing effector complexes. In this study, we show that the 122-kDa replicase subunit (p122) of crucifer-infecting Tobacco mosaic virus (cr-TMV) is a potent silencing-suppressor protein. We found that the p122 protein preferentially binds to double-stranded 21-nucleotide (nt) siRNA and microRNA (miRNA) intermediates with 2-nt 3′ overhangs inhibiting the incorporation of siRNA and miRNA into silencing-related complexes (e.g., RNA-induced silencing complex [RISC]) both in vitro and in planta but cannot interfere with previously programmed RISCs. In addition, our results also suggest that the virus infection and/or sequestration of the siRNA and miRNA molecules by p122 enhances miRNA accumulation despite preventing its methylation. However, the p122 silencing suppressor does not prevent the methylation of certain miRNAs in hst-15 mutants, in which the nuclear export of miRNAs is compromised.

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