Identification of leukemia-specific fusion gene transcripts with a novel oligonucleotide array.
BACKGROUND
Identification of specific chromosomal translocations is essential for the diagnosis and prognosis of leukemia. In this study, we employ DNA microarray technology to detect chromosomal aberrations in patients with chronic myeloid leukemia (CML) and acute myeloid leukemia (AML), as well as in leukemic cell lines.


METHODS
Reverse transcription using a random 9-mer primer was performed with total RNA from patients and leukemic cells lines. Multiplex PCR reactions using four groups of primer sets were then performed for amplification of cDNA from reverse-transcribed total RNA samples. Normal and fusion sequences were distinguished by hybridization of the amplified cDNA to a selective oligonucleotide array (SOA) containing 20-30mer synthetic probes. A total of 23 sets of oligomers were fabricated on glass slides for the detection of normal and fusion genes, as follows: BCR/ABL, AML/EAP, AML/ETO, AML/MDS, PML/RARA, NUMA1/RARA, PLZF/RARA, and CBFB/MYH.


RESULTS
Gene translocation in leukemia was effectively identified with the SOA containing various leukemia-specific fusion and normal control sequences. Leukemic fusion sequences from patients and cell lines hybridized specifically to their complementary probes. The probe sets differing by approximately 50% at their 5' or 3' ends could distinguish between normal and fusion sequences. The entire process of detection was completed within 8 hours using the SOA method.


CONCLUSIONS
Probe sets on SOA can effectively discriminate between leukemia-specific fusion and normal sequences with a chip hybridization procedure. The oligonucleotide array presents several advantages in identifying leukemic gene translocations, such as multiplex screening, relatively low cost, and speed.
Chun
Sung-Min
SM
Life Sciences Division, Korea Institute of Science and Technology, Seoul, Korea.
Kim
Yoo-Li
YL
Choi
Hee Baeg
HB
Oh
Yong-Taek
YT
Kim
Yoo-Jin
YJ
Lee
Seok
S
Kim
Tai-Gyu
TG
Yang
Eun Gyeong
EG
Park
Yong-Keun
YK
Kim
Dong-Wook
DW
Han
Byoung-Don
BD
D045744:Cell Line, Tumor; D002884:Chromosomes, Human, Pair 15; D002891:Chromosomes, Human, Pair 21; D018076:DNA, Complementary; D004273:DNA, Neoplasm / Q000235:genetics; D004277:DNA, Single-Stranded / Q000235:genetics; D050939:Gene Fusion; D006801:Humans; D020014:K562 Cells; D007938:Leukemia / Q000235:genetics*; D020411:Oligonucleotide Array Sequence Analysis; D012334:RNA, Neoplasm / Q000235:genetics; D014158:Transcription, Genetic; D014178:Translocation, Genetic
D018076:DNA, Complementary; D004273:DNA, Neoplasm; D004277:DNA, Single-Stranded; D012334:RNA, Neoplasm