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Biomedical subjects

U Linz

Publications and source records attributed to U Linz.

9 recordsLinked to original sources
PubMed1990

[Polymerase chain reaction: an overview].

In an unprecedented short time, the polymerase chain reaction (PCR) has developed into an indispensible analytical tool which facilitates gene analyses and recombinant techniques, enables DNA detection from single cells, and permits sequence determinations even from extinct species. Like the PCR process itself, the number of laboratories making use of this novel technique has been growing exponentially. This report covers critically the PCR from the basic principle to selected examples. It illustrates the strengths of the method: speed, sensitivity, and versatility and discusses aspects which still need improvement and development.

DNA↗
PubMed1990

Thermocycler temperature variation invalidates PCR results.

Among the factors which have to be optimized to ensure highest sensitivity and specificity of the polymerase chain reaction (PCR) is the processor which enables the automatic performance of the PCR assay. It has to guarantee temperature homogeneity for all samples of an individual run and run-to-run comparability. The following report illustrates that the present generation of PCR processors does not yet fulfill this requirement to satisfaction. It discusses examples of significant performance variations and their consequences for the outcome of the amplification reaction.

Biotechnology↗
PubMed1990

Systematic studies on parameters influencing the performance of the polymerase chain reaction.

To obtain a detailed understanding of the various factors influencing the polymerase chain reaction, we optimized parameters such as ion concentration, pH, primer sequence and concentration, hybridization stringency, cycle numbers, etc. Using 3 different plasmids (2 HIV2 clones and pUC18) and several genomic DNA samples from HIV-positive patients as templates, together with 6 sets of primers, the following optimal conditions were found: the DNA should be linearized, the primer concentration should be 0.1-0.2 mumol/l, and the magnesium ion concentration should be less than 2 mmol/l. The pH of the reaction mixture should be 8.5-9.0. Twenty five cycles are sufficient. For fragments greater than 10(3) bases the elongation time should be 5 min. The elongation temperature is not critical and can vary between 50 and 70 degrees C. The hybridization temperature can be used to control the specificity of the polymerase chain reaction and, finally, mismatches at the 3' end of the primer can totally inhibit the amplification.

Base Sequence↗
PubMed1988

[Warts and viruses].

The discovery that papillomaviruses are correlated with various types of cancer in man has led to increased interest in this virus group. A more detailed study of these viruses has only become possible with the introduction of recombinant DNA techniques. The most thoroughly studied representative, the bovine papillomavirus type 1, has now itself become a tool in genetic engineering.

Animals↗
PubMed1988

Promoters of bovine papillomavirus type 1: in vitro activity and utilization.

Five of six bovine papillomavirus type 1 (BPV-1) promoters which were previously mapped by determining the 5' termini of viral mRNAs from bovine fibropapillomas and BPV-1-transformed cells were found to be active under in vitro transcription conditions. Transcription initiation at each of these promoters was accurate at the nucleotide level as determined by primer extension analysis. The most active promoter in vitro was P89, a typical RNA polymerase II promoter with both TATA and CAAT boxes. The promoters P7185 and P2443 also have TATA-like boxes, but none of the other promoters contain these typical regulatory elements.

Bovine papillomavirus 1↗
PubMed1985

Chemical and biological consequences of the radioactive decay of iodine-125 in plasmid DNA.

Doubly labeled [U-14C, 5-125I]iododeoxycytidine (IdC) triphosphate was synthesized and incorporated enzymatically into defined positions of the plasmid pBR322. After storage under various conditions, the stable end products were analyzed using radio-GC, radio-HPLC, and electron microscopy. In addition, solutions of 14C-IdC-labeled DNA containing Na125I as an internal radiation source were studied to investigate the influence of internal radiolysis. Transmutation of the covalently bound 125I leads to complete destruction of the labeled nucleotide, giving rise to 14CO2 and 14CO as major products. Fragmentation of the pyrimidine base is independent of solvent and DNA configuration. Internal radiolysis caused by Na125I leads to only minor damage. Electron microscopy studies reveal that decay-induced double strand breaks (dsb) occur both at the site of decay and in areas as far as hundreds of base pairs apart from that site. Number and distribution of the breaks is strongly dependent on solvent and DNA configuration. A direct correlation exists between the extent of fragmentation of the nucleotide and the mean number of dsb.

Bromodeoxycytidine↗