Reverse Transcription with Random Pentadecamer Primers Improves the Detection Limit of a Quantitative PCR Assay for BCR-ABL Transcripts in Chronic Myeloid Leukemia: Implications for Defining Sensitivity in Minimal Residual Disease

摘要: Background: Real-time quantitative reverse transcription PCR (RQ-PCR) assay for BCR-ABL is used to monitor treatment response in chronic myeloid leukemia (CML). transcript levels decline over several years of imatinib treatment, and increasing numbers patients have transcripts at or below the limit detection. More sensitive methods are required assess whether these a long-term continuing residual disease. Methods: We random pentadecamer (R15) primers RQ-PCR compared results with our established method that uses hexamers. An increase sensitivity would be detected as an number transcripts. Results: increased by 86% R15 primers. retest 19 samples from selected CML who had no recently detectable hexamer 68% samples. Use showed variable increases control genes BCR (breakpoint cluster region), ABL1 (c-abl oncogene 1, receptor tyrosine kinase), GUSB (glucuronidase, beta), depending on gene examined. The reported /control ratio was affected, estimated detection assay, which based copy number, different each gene. Conclusions: This simple modification methodology improved transcripts. In field CML, important implications defining when undetectable. Random may also useful other assays abundance target RNA low.