ID | 31305 |
JaLCDOI | |
FullText URL | |
Author |
Inoue, Seiichi
Okamoto, Osamu
Murakami, Hiroki
Isizu, Hideo
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Abstract | A sensitive method of HLA-DRB1 typing was devised using a semi-nested polymerase chain reaction (PCR) followed by a restriction fragment length polymorphism (RFLP) analysis (semi-nested PCR-RFLP method). The first-round amplification (30 cycles) of the semi-nested PCR was performed using DRB generic primer pairs and the second round of PCRs (20 cycles) were performed using DRB1 group-specific primers. The products of the second round PCRs were digested with restriction endonucleases for the typing of HLA-DRB1 alleles. By this method, HLA-DRB1 typing was possible from 10 pg of genomic DNA extracted from lymphocytes and from 0.5 microliter of 1,000 times diluted blood without DNA extraction. HLA-DRB1 alleles could be typed from a 2-mm long bloodstained cotton thread prepared from 10 times diluted blood and from a 2-mm thread of whole blood bloodstains stored at room temperature for 2 years. From the mixture of blood of two individuals with different genotypes, DRB1 alleles of the minor component were detected down to 1/1,000 of the major component. This semi-nested PCR-RFLP method is useful for HLA-DRB1 typing from extremely small amounts of DNA and from mixed samples. |
Keywords | polymorphism
HLA-DRB1
polymerase chain reaction
dsmi-nested PCR
restricton fragment length polymotphism
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Amo Type | Article
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Publication Title |
Acta Medica Okayama
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Published Date | 1998-12
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Volume | volume52
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Issue | issue6
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Publisher | Okayama University Medical School
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Start Page | 289
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End Page | 296
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ISSN | 0386-300X
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NCID | AA00508441
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Content Type |
Journal Article
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language |
English
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File Version | publisher
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Refereed |
True
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PubMed ID | |
Web of Science KeyUT |