[Using real-time polymerase chain reaction to quantitate bcr-abl mRNA].
OBJECTIVE
When leukemia patients achieve complete remission after chemotherapy, a few tumor cells still exist in other tissues outside bone marrow, which is called minimal residual disease (MRD), and it is the base of relapse. To cure leukemia, we should not only find MRD in time, but also quantitate MRD for instructing treatment and predicting prognosis. This study was to establish a real-time reverse transcription-polymerase chain reaction (RT-PCR) system to quantitate bcr-abl mRNA.


METHODS
bcr-abl mRNA in leukemia cell line K562 was amplified by RT-PCR. T-A clone was used to construct the combined plasmid to be standard template; the standard curve of bcr-abl oncogene was drawn. bcr-abl mRNA in bone marrow samples of 16 chronic myelocytic leukemia (CML) patients was quantitated by this method. The sensitivity, stability, and repetition of this method were evaluated.


RESULTS
The sensitivity is 10 copies of the recombined plasmid. The coefficient variations (CV) of repetition and stability were 2.19% and 3.21%. The correlation (R) of the standard curve was 0.984. The median bcr-abl level of the 16 CML patients was 4.58x10(4) kb/microg RNA.


CONCLUSIONS
Real-time PCR has high sensitivity, repetition, and specificity. It can quantitate the copy number of bcr-abl oncogene.
Liu
Jin-Xiang
JX
Department of Hematology, The First Affiliated Hospital, Jilin University, Changchun, Jilin, 130021, P. R. China.
Liu
Jun-Li
JL
Wang
Guan-Jun
GJ
D001853:Bone Marrow / Q000378:metabolism / Q000473:pathology; D016044:Fusion Proteins, bcr-abl / Q000096:biosynthesis* / Q000235:genetics; D018922:HL-60 Cells; D006801:Humans; D020014:K562 Cells; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D012334:RNA, Neoplasm / Q000235:genetics; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
D012333:RNA, Messenger; D012334:RNA, Neoplasm; D016044:Fusion Proteins, bcr-abl