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Titolo:
Quantitative detection of single nucleotide polymorphisms for a pooled sample by a bioluminometric assay coupled with modified primer extension reactions (BAMPER)
Autore:
Zhou, GH; Kamahori, M; Okano, K; Chuan, G; Harada, K; Kambara, H;
Indirizzi:
Hitachi Ltd, Cent Res Lab, Kokubunji, Tokyo 1858601, Japan Hitachi Ltd Kokubunji Tokyo Japan 1858601 Kokubunji, Tokyo 1858601, Japan
Titolo Testata:
NUCLEIC ACIDS RESEARCH
fascicolo: 19, volume: 29, anno: 2001,
pagine: NIL_33 - NIL_43
SICI:
0305-1048(20011001)29:19<NIL_33:QDOSNP>2.0.ZU;2-C
Fonte:
ISI
Lingua:
ENG
Soggetto:
REAL-TIME; POINT MUTATIONS; DNA; PCR; DISCRIMINATION; IDENTIFICATION; PYROPHOSPHATE; PERFORMANCE;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
33
Recensione:
Indirizzi per estratti:
Indirizzo: Kambara, H Hitachi Ltd, Cent Res Lab, 1-280 Higashi Koigakubo, Kokubunji, Tokyo 1858601, Japan Hitachi Ltd 1-280 Higashi Koigakubo Kokubunji Tokyo Japan 1858601
Citazione:
G.H. Zhou et al., "Quantitative detection of single nucleotide polymorphisms for a pooled sample by a bioluminometric assay coupled with modified primer extension reactions (BAMPER)", NUCL ACID R, 29(19), 2001, pp. NIL_33-NIL_43

Abstract

A now method for SNP analysis based on the detection of pyrophosphate (PPI) Is demonstrated, which is capable of detecting small allele frequency differences between two DNA pools for genetic association studies other than SNP typing. The method Is based on specific primer extension reactions coupled with PPI detection. As the specificity of the primer-directed extension Is not enough for quantitative SNP analysis, artificial mismatched bases are Introduced Into the 3 ' -terminal regions of the specific primers as a way of Improving the switching characteristics of the primer extension reactions. The best position In the primer for such artificial mismatched bases Is the third position from the primer 3 ' -terminus. Contamination with endogenous PPI, which produces a large background signal level In SNP analysis,was removed using PPase to degrade the PPl during the sample preparation process. It Is possible to accurately and quantitatively analyze SNPs using a set of primers that correspond to the wild-type and mutant DNA segments. The termini of these primers are at the mutation positions. Various types of SNPs were successfully analyzed. It was possible to very accurately determine SNPs with frequencies as low 0.02. It Is very reproducible and the allele frequency difference can be determined. it Is accurate enough to detectmeaningful genetic differences among pooled DNA samples. The method Is sensitive enough to detect 14 amol ssM13 DNA. The proposed method seems very promising In terms of realizing a cost-effective, large-scale human genetic testing system.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 18/01/20 alle ore 13:43:39