Real-time reverse transcription loop-mediated isothermal amplification for rapid and simple quantification of WT1 mRNA.

Abstract

OBJECTIVES:

This study developed a novel MRD monitoring method targeting Wilms' tumor gene (WT1) mRNA using reverse transcription loop-mediated isothermal amplification (RT-LAMP).

DESIGN AND METHODS:

A primer set for the assay was designed on the basis of the sequences between the 17AA and KTS regions of WT1mRNA. WT1 mRNA was quantified by real-time RT-LAMP and the accuracy of RT-LAMP was compared with that of real-time RT-PCR.

RESULTS:

The standard curve was expressed as a linear relationship between the log copy numbers of WT1 mRNA ranging from 6.8 x 10 to 6.8 x 10(9) copies and the threshold time with a correlation coefficient of R(2) > 0.994. The measured values obtained by RT-LAMP strongly correlated with those obtained by real-time RT-PCR.

CONCLUSION:

RT-LAMP can be used to determine WT1 mRNA expression levels. This assay will contribute to a more specific, simple, and rapid MRD monitoring than conventional assays.

• Authors: Soji Morishita, Hidenori Tani, Shinya Kurata, Kazunori Nakamura, Satoshi Tsuneda, Yuji Sekiguchi, Naohiro Noda
• Journal: Clinical biochemistry (Clin Biochem) Vol. 42 Issue 6 Pg. 515-20 (Apr 2009) ISSN: 1873-2933 [Electronic] United States
• PMID: 19297684 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• DNA Primers
• RNA, Messenger

Topics

• Base Sequence
• DNA Primers
• Gene Expression
• Genes, Wilms Tumor
• Humans
• Leukemia (diagnosis, genetics)
• Molecular Sequence Data
• Neoplasm, Residual
• Prognosis
• RNA, Messenger (analysis)
• Reverse Transcriptase Polymerase Chain Reaction (methods)
• Sensitivity and Specificity