Enrichment of mutated K-RAS from human stool DNA by a combined magnetic capture hybridization and PCR-RFLP assay.

Publikation: Beiträge in SammelwerkenAbstracts in KonferenzbändenForschungbegutachtet

Standard

Enrichment of mutated K-RAS from human stool DNA by a combined magnetic capture hybridization and PCR-RFLP assay. / Schneider, Mandy; Steinberg, Pablo; Scholtka, Bettina.
Wissen teilen - Chancen nutzen: 28. Deutscher Krebskongress, Berlin, Februar 2008 und 3. Krebsaktionstag, Berlin, Februar 2008: Abstracts . Hrsg. / M. Kaufmann; A. Rody; M. Bamberg. Band 31 Karger Verlag für Medizin und Naturwissenschaften , 2008. S. 59 (Onkologie; Band 31, Nr. Suppl. 1).

Publikation: Beiträge in SammelwerkenAbstracts in KonferenzbändenForschungbegutachtet

Harvard

Schneider, M, Steinberg, P & Scholtka, B 2008, Enrichment of mutated K-RAS from human stool DNA by a combined magnetic capture hybridization and PCR-RFLP assay. in M Kaufmann, A Rody & M Bamberg (Hrsg.), Wissen teilen - Chancen nutzen: 28. Deutscher Krebskongress, Berlin, Februar 2008 und 3. Krebsaktionstag, Berlin, Februar 2008: Abstracts . Bd. 31, Onkologie, Nr. Suppl. 1, Bd. 31, Karger Verlag für Medizin und Naturwissenschaften , S. 59, 28. Deutscher Krebskongress - 2008, Berlin, Deutschland, 20.02.08. https://doi.org/10.1159/000115424

APA

Schneider, M., Steinberg, P., & Scholtka, B. (2008). Enrichment of mutated K-RAS from human stool DNA by a combined magnetic capture hybridization and PCR-RFLP assay. In M. Kaufmann, A. Rody, & M. Bamberg (Hrsg.), Wissen teilen - Chancen nutzen: 28. Deutscher Krebskongress, Berlin, Februar 2008 und 3. Krebsaktionstag, Berlin, Februar 2008: Abstracts (Band 31, S. 59). (Onkologie; Band 31, Nr. Suppl. 1). Karger Verlag für Medizin und Naturwissenschaften . https://doi.org/10.1159/000115424

Vancouver

Schneider M, Steinberg P, Scholtka B. Enrichment of mutated K-RAS from human stool DNA by a combined magnetic capture hybridization and PCR-RFLP assay. in Kaufmann M, Rody A, Bamberg M, Hrsg., Wissen teilen - Chancen nutzen: 28. Deutscher Krebskongress, Berlin, Februar 2008 und 3. Krebsaktionstag, Berlin, Februar 2008: Abstracts . Band 31. Karger Verlag für Medizin und Naturwissenschaften . 2008. S. 59. (Onkologie; Suppl. 1). doi: 10.1159/000115424

Bibtex

@inbook{60689a172e4d412ab1ad2cb63ed4bf2f,
title = "Enrichment of mutated K-RAS from human stool DNA by a combined magnetic capture hybridization and PCR-RFLP assay.",
abstract = "Introduction: Although many techniques for DNA mutation analysis areavailable, most of them lack the sensitivity to detect alterations in human stoolDNA at a comparable level to that present in tissues. In faeces usually onemutated DNA molecule among hundred to ten thousand wild-type DNA mol-ecules does in fact exist. The aim of the project was to develop an assay toenrich the mutated K-RAS from a mixture of wild-type and mutant DNA instool and to compare the achieved detection rate with that of an establishedPCR-RFLP method.Methods: A combined magnetic capture hybridization and PCR-RFLP assayto detect K-RAS codon 12 or 13 mutations was developed. Briefly, two PCRreactions with an intermediate restriction digest followed by a hybridizationreaction with a biotinylated single-strand DNA probe were performed.Positive controls were derived from human carcinoma cell lines. Serial dilu-tions of mutant and wild-type controls were prepared to determine the sensi-tivity of the assay. Furthermore, a few stool samples and tissues from patientswith colon or rectum carcinomas or prestages were analyzed. All observedmutations were confirmed by sequencing.Results: The combination of restriction digest and magnetic capturehybridization led to a 100-fold increased detection limit for K-RAS mutationsin both, codons 12 and 13, when compared to the conventional PCR-RFLPassay. With the conventional PCR-RFLP assay it was not possible to detectalterations in stool samples of patients with known K-RAS gene mutations intheir corresponding preneoplastic or neoplastic intestinal tissues. In contrast,by using the combined magnetic capture hybridization and PCR-RFLP assayK-RAS mutations could be found in stool samples as previously observed inthe corresponding biopsies.Conclusion: The combined magnetic capture hybridization and PCR-RFLPassay leads to an enrichment of mutated K-RAS and is sensitive enough todetect K-RAS codon 12 or 13 mutations in stool DNA from patients with pre-neoplastic and neoplastic intestinal lesions",
keywords = "Biology, PCR, DNA",
author = "Mandy Schneider and Pablo Steinberg and Bettina Scholtka",
note = "Ersch. als Zeitschriftensonderheft: Onkologie 2008;31(suppl 1):1–211 Abstracts; 28. Deutscher Krebskongress - 2008 ; Conference date: 20-02-2008 Through 23-02-2008",
year = "2008",
month = jan,
day = "1",
doi = "10.1159/000115424",
language = "English",
isbn = "978-3-8055-8536-1 ",
volume = "31",
series = "Onkologie",
publisher = "Karger Verlag f{\"u}r Medizin und Naturwissenschaften ",
number = "Suppl. 1",
pages = "59",
editor = "M. Kaufmann and A. Rody and M. Bamberg",
booktitle = "Wissen teilen - Chancen nutzen",
address = "Switzerland",

}

RIS

TY - CHAP

T1 - Enrichment of mutated K-RAS from human stool DNA by a combined magnetic capture hybridization and PCR-RFLP assay.

AU - Schneider, Mandy

AU - Steinberg, Pablo

AU - Scholtka, Bettina

N1 - Conference code: 28

PY - 2008/1/1

Y1 - 2008/1/1

N2 - Introduction: Although many techniques for DNA mutation analysis areavailable, most of them lack the sensitivity to detect alterations in human stoolDNA at a comparable level to that present in tissues. In faeces usually onemutated DNA molecule among hundred to ten thousand wild-type DNA mol-ecules does in fact exist. The aim of the project was to develop an assay toenrich the mutated K-RAS from a mixture of wild-type and mutant DNA instool and to compare the achieved detection rate with that of an establishedPCR-RFLP method.Methods: A combined magnetic capture hybridization and PCR-RFLP assayto detect K-RAS codon 12 or 13 mutations was developed. Briefly, two PCRreactions with an intermediate restriction digest followed by a hybridizationreaction with a biotinylated single-strand DNA probe were performed.Positive controls were derived from human carcinoma cell lines. Serial dilu-tions of mutant and wild-type controls were prepared to determine the sensi-tivity of the assay. Furthermore, a few stool samples and tissues from patientswith colon or rectum carcinomas or prestages were analyzed. All observedmutations were confirmed by sequencing.Results: The combination of restriction digest and magnetic capturehybridization led to a 100-fold increased detection limit for K-RAS mutationsin both, codons 12 and 13, when compared to the conventional PCR-RFLPassay. With the conventional PCR-RFLP assay it was not possible to detectalterations in stool samples of patients with known K-RAS gene mutations intheir corresponding preneoplastic or neoplastic intestinal tissues. In contrast,by using the combined magnetic capture hybridization and PCR-RFLP assayK-RAS mutations could be found in stool samples as previously observed inthe corresponding biopsies.Conclusion: The combined magnetic capture hybridization and PCR-RFLPassay leads to an enrichment of mutated K-RAS and is sensitive enough todetect K-RAS codon 12 or 13 mutations in stool DNA from patients with pre-neoplastic and neoplastic intestinal lesions

AB - Introduction: Although many techniques for DNA mutation analysis areavailable, most of them lack the sensitivity to detect alterations in human stoolDNA at a comparable level to that present in tissues. In faeces usually onemutated DNA molecule among hundred to ten thousand wild-type DNA mol-ecules does in fact exist. The aim of the project was to develop an assay toenrich the mutated K-RAS from a mixture of wild-type and mutant DNA instool and to compare the achieved detection rate with that of an establishedPCR-RFLP method.Methods: A combined magnetic capture hybridization and PCR-RFLP assayto detect K-RAS codon 12 or 13 mutations was developed. Briefly, two PCRreactions with an intermediate restriction digest followed by a hybridizationreaction with a biotinylated single-strand DNA probe were performed.Positive controls were derived from human carcinoma cell lines. Serial dilu-tions of mutant and wild-type controls were prepared to determine the sensi-tivity of the assay. Furthermore, a few stool samples and tissues from patientswith colon or rectum carcinomas or prestages were analyzed. All observedmutations were confirmed by sequencing.Results: The combination of restriction digest and magnetic capturehybridization led to a 100-fold increased detection limit for K-RAS mutationsin both, codons 12 and 13, when compared to the conventional PCR-RFLPassay. With the conventional PCR-RFLP assay it was not possible to detectalterations in stool samples of patients with known K-RAS gene mutations intheir corresponding preneoplastic or neoplastic intestinal tissues. In contrast,by using the combined magnetic capture hybridization and PCR-RFLP assayK-RAS mutations could be found in stool samples as previously observed inthe corresponding biopsies.Conclusion: The combined magnetic capture hybridization and PCR-RFLPassay leads to an enrichment of mutated K-RAS and is sensitive enough todetect K-RAS codon 12 or 13 mutations in stool DNA from patients with pre-neoplastic and neoplastic intestinal lesions

KW - Biology

KW - PCR

KW - DNA

U2 - 10.1159/000115424

DO - 10.1159/000115424

M3 - Published abstract in conference proceedings

SN - 978-3-8055-8536-1

VL - 31

T3 - Onkologie

SP - 59

BT - Wissen teilen - Chancen nutzen

A2 - Kaufmann, M.

A2 - Rody, A.

A2 - Bamberg, M.

PB - Karger Verlag für Medizin und Naturwissenschaften

T2 - 28. Deutscher Krebskongress - 2008

Y2 - 20 February 2008 through 23 February 2008

ER -

DOI