An optimized multiplex polymerase chain reaction (PCR) for detection of BCR-ABL fusion mRNAs in haematological disorders.
A rapid and simple polymerase chain reaction (PCR) method is described that is capable of identifying any of the BCR-ABL transcripts that have yet been described in chronic myeloid or acute lymphoblastic leukaemia. Randomly primed cDNA is synthesized from leucocyte RNA and amplified in a single reaction containing four oligonucleotide primers (multiplex PCR). Different size products are generated from ela2 (p190) and b3a2 or b2a2 (p210) BCR-ABL transcripts which are readily and unambiguously distinguishable after agarose gel electrophoresis without the need for either nested PCR or hybridization. Chronic myeloid leukaemia cells are readily detectable even when diluted 1 in 1000 with normal blood. Samples which do not have BCR-ABL rearrangements produce a single band derived from the normal BCR gene, and the presence of this band controls for adequate RNA and cDNA preparation. Using this assay we have detected BCR-ABL transcripts in a variety of haematological disorders.
Cross
N C
NC
LRF Centre for Adult Leukaemia, Royal Postgraduate Medical School, London, UK.
Melo
J V
JV
Feng
L
L
Goldman
J M
JM
D001483:Base Sequence; D018076:DNA, Complementary / Q000032:analysis / Q000096:biosynthesis / Q000235:genetics; D004587:Electrophoresis, Agar Gel; D016044:Fusion Proteins, bcr-abl / Q000235:genetics*; D005784:Gene Amplification; D006402:Hematologic Diseases / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D012333:RNA, Messenger / Q000032:analysis* / Q000235:genetics; D014158:Transcription, Genetic / Q000235:genetics
D018076:DNA, Complementary; D012333:RNA, Messenger; D016044:Fusion Proteins, bcr-abl