An improved fluorescence polarization assay in 5'-nuclease reaction for gene promoter methylation detection.

Abstract:

:The detection of gene promoter methylation plays increasing roles in personalized medicine. In this study, an improved gene promoter methylation assay based on fluorescence polarization in 5'-nuclease reaction was developed. The novel assay offered a homogeneous annealing and cleavage reaction fully integrated with PCR which used a probe labeled with fluorescence without quencher to obtain the decreased fluorescence polarization values. In this platform, gene promoter methylated and unmethylated alleles were detected simultaneously in a tube. O(6)-methylguanine-DNA methyltransferase gene promoter methylation in 103 glioma tissue samples and epidermal growth factor receptor gene promoter methylation in 116 primary non-small-cell lung carcinoma tissue samples were detected by the novel assay and sequencing, absolute quantitative analysis of methylated allele in parallel. The accuracy of the results measured by the improved fluorescence polarization assay was evaluated using the paired-samples t test. No significant difference was found ( P>0.05). Therefore, the improved fluorescence polarization assay in 5'-nuclease reaction demonstrated a homogeneous, reliable and cost-effective method for gene promoter methylation analysis in clinic. That would provide a scientific basis for applying a reasonable therapeutic regimen in future treatment.

journal_name

J Biotechnol

journal_title

Journal of biotechnology

authors

Su H,Wu Z,Wang Y,Jiang Y,Qiang S,Cheng H,Liu W,Zhang J

doi

10.1016/j.jbiotec.2015.07.003

subject

Has Abstract

pub_date

2015-10-10 00:00:00

pages

81-6

eissn

0168-1656

issn

1873-4863

pii

S0168-1656(15)30060-2

journal_volume

211

pub_type

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