New primers for methylation-specific polymerase chain reaction enhance specificity of detecting STAT1 methylation
Journal
Taiwanese Journal of Obstetrics and Gynecology
Journal Volume
51
Journal Issue
1
Pages
43-49
Date Issued
2012
Author(s)
Abstract
Objective: Signal transducer and activator of transcription (STAT)1 is a key tumor suppressor, which is always methylated in a variety of human cancers. However, nonspecific primers for the detection of specific promoter hypermethylation of STAT1 gene can lead to false-positive or false-negative results for gene methylation. Materials and Methods: We designed new primers for the detection of STAT1 methylation and compared the sensitivities and specificities of these new primers with prior published primers by methylation-specific polymerase chain reaction (PCR) from ovarian clear cell carcinomas. The mRNA expression levels of STAT1 in these cancerous tissues were also evaluated by reverse-transcriptase PCR and correlated with the results of promoter methylation of STAT1 gene. Results: Nine (39%) of the 23 samples detected by the new primers and 13 samples (56%) detected by prior published primers showed STAT1 methylation. A direct DNA sequencing test revealed that four of the 13 samples (30.8%) showed false positivity for STAT1 methylation using the prior published primers. In contrast, none of the nine samples was false-positive for the detection of STAT1 methylation using the new primers. The new primers for the detection of STAT1 methylation showed 100% specificity and 100% sensitivity without false positivity. Conclusion: Specific primers for methylation-specific PCR are mandatory for the accurate detection of STAT1 gene methylation. Besides, specific primers can generate correct interpretation of STAT1 gene methylation, and its correlation with the clinicopathological characteristics and outcome of cancer patients. ? 2012.
SDGs
Other Subjects
messenger RNA; primer DNA; STAT1 protein; article; cancer tissue; clear cell carcinoma; controlled study; CpG island; DNA methylation; DNA sequence; false negative result; false positive result; female; gene expression; human; human tissue; intermethod comparison; methylation specific polymerase chain reaction; nucleotide sequence; ovary carcinoma; promoter region; reverse transcription polymerase chain reaction; sensitivity and specificity; Adenocarcinoma, Clear Cell; DNA Methylation; DNA Primers; False Positive Reactions; Female; Humans; Ovarian Neoplasms; Polymerase Chain Reaction; Promoter Regions, Genetic; RNA, Messenger; Sensitivity and Specificity; STAT1 Transcription Factor
Type
journal article