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M94A0080.TXT
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1994-10-01
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Document 0080
DOCN M94A0080
TI Capillary electrophoresis: detection of hybridization between synthetic
oligonucleotides and HIV-1 genomic DNA amplified by polymerase-chain
reaction.
DT 9412
AU Bianchi N; Mischiati C; Feriotto G; Fiorentino D; Di Biase S; Apicella
N; Gambari R; Biochemistry Institute, Ferrara University, Italy.
SO J Virol Methods. 1994 May;47(3):321-9. Unique Identifier : AIDSLINE
MED/94351052
AB The polymerase chain reaction (PCR) is one of the most efficient
techniques for measuring the viral load of HIV-infected samples.
Determination of the specificity of PCR products is usually based on
Southern blotting and hybridization of the amplified DNA to radioactive
oligonucleotide probes specific for sequences comprised between the PCR
primers. The recent introduction of capillary electrophoresis (CE) for
identification of HIV-1 and HTLV-I PCR products appears interesting in
light of its reproducibility, sensitivity and because it is fast and
suitable for detection of DNA/DNA and DNA/RNA hybrids. We demonstrate
that specific hybridization of a HIV-1 oligonucleotide probe to
single-stranded DNA obtained by unbalanced PCR is detectable by
capillary electrophoresis. This enabled us the application of a
one-step, non-radioactive protocol to demonstrate the specificity of
amplification of HIV-1 genomic sequences by PCR. This procedure is
simple, reproducible and is suggested as an integral part of automated
diagnostic systems based on the use of laboratory work stations for DNA
isolation, preparation of PCR reactions and analysis of PCR products.
DE Base Sequence DNA, Viral/*CHEMISTRY/ISOLATION & PURIF
Electrophoresis/*METHODS Genome, Viral HIV-1/*GENETICS Molecular
Sequence Data Nucleic Acid Hybridization Oligonucleotides/*CHEMISTRY
Polymerase Chain Reaction Support, Non-U.S. Gov't JOURNAL ARTICLE
SOURCE: National Library of Medicine. NOTICE: This material may be
protected by Copyright Law (Title 17, U.S.Code).