Real-time reverse transcription loop-mediated isothermal amplification for rapid and simple quantification of WT1 mRNA.
OBJECTIVE
This study developed a novel MRD monitoring method targeting Wilms' tumor gene (WT1) mRNA using reverse transcription loop-mediated isothermal amplification (RT-LAMP).


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.


CONCLUSIONS
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.
Morishita
Soji
S
Department of Life Science and Medical Bio-science, Waseda University, 2-2 Wakamatsu-cho, Shinjuku-ku, Tokyo, Japan.
Tani
Hidenori
H
Kurata
Shinya
S
Nakamura
Kazunori
K
Tsuneda
Satoshi
S
Sekiguchi
Yuji
Y
Noda
Naohiro
N
D001483:Base Sequence; D017931:DNA Primers; D015870:Gene Expression; D016162:Genes, Wilms Tumor; D006801:Humans; D007938:Leukemia / Q000175:diagnosis* / Q000235:genetics; D008969:Molecular Sequence Data; D018365:Neoplasm, Residual; D011379:Prognosis; D012333:RNA, Messenger / Q000032:analysis*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
D017931:DNA Primers; D012333:RNA, Messenger