A simple and robust real-time qPCR method for the detection of PIK3CA mutations.
Sci Rep
; 8(1): 4290, 2018 03 09.
Article
en En
| MEDLINE
| ID: mdl-29523855
ABSTRACT
PIK3CA mutations are seemingly the most common driver mutations in breast cancer with H1047R and E545K being the most common of these, accounting together for around 60% of all PIK3CA mutations and have promising therapeutic implications. Given the low sensitivity and the high cost of current genotyping methods we sought to develop fast, simple and inexpensive assays for PIK3CA H1047R and E545K mutation screening in clinical material. The methods we describe are based on a real-time PCR including a mutation specific primer combined with a non-productive oligonucleotide which inhibits wild-type amplification and a parallel internal control reaction. We demonstrate consistent detection of PIK3CA H1047R mutant DNA in genomic DNA extracted from frozen breast cancer biopsies, FFPE material or cancer cell lines with a detection sensitivity of approximately 5% mutant allele fraction and validate these results using both Sanger sequencing and deep next generation sequencing methods. The detection sensitivity for PIK3CA E545K mutation was approximately 10%. We propose these methods as simple, fast and inexpensive diagnostic tools to determine PIK3CA mutation status.
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Neoplasias de la Mama
/
Pruebas Genéticas
/
Fosfatidilinositol 3-Quinasa Clase I
/
Reacción en Cadena en Tiempo Real de la Polimerasa
/
Mutación
Tipo de estudio:
Diagnostic_studies
/
Prognostic_studies
Límite:
Female
/
Humans
Idioma:
En
Revista:
Sci Rep
Año:
2018
Tipo del documento:
Article
País de afiliación:
Reino Unido