Accurate and Rapid Detection of Heterozygous Carriers of a Deletion by Combined Polymerase Chain Reaction and High-Performance Liquid Chromatography

We have developed a technique to detect accurately heterozygous carriers of a deletion. Specific target sequences were amplified by the polymerase chain reaction (PCR), and the products subsequently were analyzed by high-performance liquid chromatography. Examples from four loci demonstrated that 24...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 1992-10, Vol.89 (19), p.9126-9130
Main Authors: Asakawa, Jun-Ichi, Satoh, Chiyoko, Yamasaki, Yosuke, Chen, Shi-Han
Format: Article
Language:English
Subjects:
Alleles
Base Sequence
carriers
Chromatography, High Pressure Liquid - methods
Deoxyribonucleic acid
detection
DNA
DNA - genetics
Duchenne muscular dystrophy
Duchenne's muscular dystrophy
Exons
Female
Gene Deletion
Genetic Carrier Screening
Genetic loci
Genetics
Genomics
Haptoglobins - genetics
Hemoglobins - genetics
Hemophilia A - genetics
Hemophilia B
heterozygosity
high-performance liquid chromatography
Homozygotes
Humans
Male
man
Molecular Sequence Data
Mothers
Oligodeoxyribonucleotides
Polymerase chain reaction
Polymerase Chain Reaction - methods
Scientific imaging
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Summary:We have developed a technique to detect accurately heterozygous carriers of a deletion. Specific target sequences were amplified by the polymerase chain reaction (PCR), and the products subsequently were analyzed by high-performance liquid chromatography. Examples from four loci demonstrated that 24-27 cycles of amplification for a single-copy DNA, based on 50 ng of genomic DNA, results in excellent quantitation that readily permits the detection of heterozygous carriers of a deletion. We have demonstrated that triplex PCR (three targets in a single PCR) entails no loss of precision. We also have demonstrated that this method can accurately differentiate the heterozygous carriers of a deletion from normal individuals in four family studies, three for Duchenne muscular dystrophy patients and one for a hemophilia B patient.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.89.19.9126