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The restriction enzyme analysis of III/2D and 6/42 strains and selected variants III/2D, III/2M, 6/42D and 6/42M selected at the presence of specific antibodies has been carried out. One additional fragment of 24 kbp was found after DNA digestion of the III/2D variant with Hind III enzyme. Electrophoretic patterns of DNA of the III/2D variant after digestion with Pst I and Eco RI enzymes revealed the lack of 6.2 kbp and 12.7 kbp fragments confirming the shifting of cleavage sites compared with parental DNA of the III/2 strain. No shifts in cleavage sites were observed in the case of Ш/2М, 6/42D and 6/42M DNAs. The data suggested that the increased resistance to nautralization with polyclonal antibody of the III/2D could result from genotype mutation.
Restriction enzyme analysis has been done for the DNA Agille and III/3 strains of the BHV 1 virus, and variants selected from these strains after neutralization with polyclonal antibodies. It has been shown that the electrophoretic pattern of DNA of the 8/-2 variant which was selected from the Agille strain and which has had higher resistance to neutralization to antibodies has been identical to the electrophoretic pattern of the Agille parent strain. This may suggest that the higher resistance to neutralization of the 8/-2 variant has been a results of phenotype variability. No difference between eletrophoretic patterns of the III/3 strain and variants III/3D and III/3H has been discovered. Digestion with Hind III, Pst I, Bam HII, Eco RI enzymes differentiate the Agille and III/3 strain.
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Application of analysis of nucleic acid using restriction enzymes in parasitology is discussed. The main advantage of the new technique is the possibility of direct and detailed studies at the level of DNA. At present, the "genetic probe" becomes more and more commonly applied to indentification of both parasites and their transmitters. It appears that the technique of restriotion analysis is of great significance for solving taxonomic problems in parasitology.
The aim of this study was to determine the subtype of bovine herpesvirus 1 (BHV1) strains currently circulating in the cattle population in Poland. Altogether 224 nasal swabs, 36 lung tissue samples and 14 samples of fluid after embryo washing in cows were tested by virus isolation test. Five field strains of BHV1 were isolated. These isolates as well as 4 archival and 2 reference strains of BHV1 were subjected to restriction enzyme analysis. Viral DNA was cleaved with Hind III, Hpa I and Pst I enzymes. After digestion with Hind III typical patterns for BHV1.1 subtype were observed in all field strains. Among archival strains 2 belonged to BHV1.1 and 2 others to B-HV1.2a subtype. These findings were confirmed by digestion with other enzymes. The presented study showed that strains of BHV1.1 subtype are dominant in cattle in Poland.
Field strains of BHV1 isolated from bovine semen at different periods in Poland have been studied by the restriction enzyme analysis of DNA. It has been shown that the Aguille strain isolated in 1976 belongs to the subtype BHV 1.2 which comprises IPV strains. But strains III/2, III/3 and 6/42 isolated in 1981 belong to the subtype BHV 1.1 which comprises IBR strains.
The aim of the study was to obtain genotypic differentiation of fungi isolated from milk samples of dairy cows showing clinical signs of mastitis. Twenty strains of fungi were isolated from milk samples and identified as Candida, and then classified into seven different species by API Candida and API ID 32C tests (bioMerieux). Next, the genomic DNA was isolated from each fungal strain and amplified with ITS1 and NL2 primers. Amplification products were digested with HpaII and EcoRI restriction enzymes, while the restriction profiles were resolved in VersaDoc imaging apparatus with the assistance of Quantity One software. PCR with ITS1 and NL2 primers produced DNA fragments of various lengths, raging from 700 to 1000 bp. Their molecular weight was dependent on the fungal species from which DNA was isolated. Comparison of restriction fragment length confirmed differences between species; however, strains similarly classified based on phenotypic characteristics also revealed differences in the restriction fragment profile. For none of the investigated species was a characteristic, uniform genetic profile obtained. Results of the presented study revealed that the examined species did not give a uniform genetic profile, suggesting that Candida strains phenotypically belonging to the same species may have varied genotypes in the analyzed restrictive places.
W identyfikacji drożdży wykorzystuje się metody klasyczne i ich modyfikacje polegające na ocenie cech fenotypowych oraz metody nowoczesne oparte na analizie polimorfizmu materiału genetycznego. Metody klasyczne są powszechnie stosowane, jednak ze względu na szybką i precyzyjną identyfikację coraz częściej wykorzystywane są techniki biologii molekularnej. W artykule opisano techniki stosowane w identyfikacji gatunkowej drożdży oparte na analizie regionów rybosomalnych (sekwencjonowanie, analiza restrykcyjna).
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