Detection of APC mutation by real-time polymerase chain reaction and melting curve analysis.
- Author:
Wei-wei ZHENG
1
;
Zhong-chen LIU
;
Qing-ge LI
Author Information
- Publication Type:Journal Article
- MeSH: Adenomatous Polyposis Coli Protein; analysis; genetics; Base Sequence; Colorectal Neoplasms; chemistry; genetics; DNA, Neoplasm; analysis; Genes, APC; Humans; Mutation; Polymerase Chain Reaction; methods; Sequence Deletion
- From: Chinese Journal of Pathology 2005;34(10):669-671
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo establish a simple, reliable and low cost approach for clinical detection of APC mutation.
METHODSUsing SYBR Green I as the real-time polymerase chain reaction (PCR) product indicator, a DNA fragment of 270 bp targeting APC_1309 mutation (5 bp deletion) was amplified from the sample DNA. A short fragment (40/35 bp) was then amplified from the 270 bp PCR product, followed by melting curve analysis from 65 degrees C to 99 degrees C at 0.5 degrees C/step.
RESULTSA total of 18 paraffin-embedded tumor samples were analyzed, of which 7 were tested positive for the mutation and 11 were negative. No mutation was detected in any of the 20 normal peripheral blood samples.
CONCLUSIONSReal-time PCR melting curve analysis can be used for routine APC mutation detection. The simple design, low cost and high reliability should allow similar applications to the analysis of a variety of other gene mutations.
