title: A pipeline with multiplex reverse transcription polymerase chain reaction and microarray for screening of chromosomal translocations in leukemia.
abstract: Chromosome rearrangements and fusion genes present major portion of leukemogenesis and contribute to leukemic subtypes. It is practical and helpful to detect the fusion genes in clinic diagnosis of leukemia. Present application of reverse transcription polymerase chain reaction (RT-PCR) method to detect the fusion gene transcripts is effective, but time- and labor-consuming. To set up a simple and rapid system, we established a method that combined multiplex RT-PCR and microarray. We selected 15 clinically most frequently observed chromosomal rearrangements generating more than 50 fusion gene variants. Chimeric reverse primers and chimeric PCR primers containing both gene-specific and universal sequences were applied in the procedure of multiplex RT-PCR, and then the PCR products hybridized with a designed microarray. With this approach, among 200 clinic samples, 63 samples were detected to have gene rearrangements. All the detected fusion genes positive and negative were validated with RT-PCR and Sanger sequencing. Our data suggested that the RT-PCR-microarray pipeline could screen 15 partner gene pairs simultaneously at the same accuracy of the fusion gene detection with regular RT-PCR. The pipeline showed effectiveness in multiple fusion genes screening in clinic samples.
authors.affiliation: School of Life Science and Technology, Tongji University, Shanghai 200092, China ; National Engineering Research Center for Biochip at Shanghai, Shanghai 201203, China.
authors.forename: Fei-Fei
authors.initials: FF
authors.lastname: Xiong
authors.forename: Ben-Shang
authors.initials: BS
authors.lastname: Li
authors.forename: Chun-Xiu
authors.initials: CX
authors.lastname: Zhang
authors.forename: Hui
authors.initials: H
authors.lastname: Xiong
authors.forename: Shu-Hong
authors.initials: SH
authors.lastname: Shen
authors.forename: Qing-Hua
authors.initials: QH
authors.lastname: Zhang
mesh_terms: D002877:Chromosomes, Human / Q000235:genetics*; D005820:Genetic Testing; D018922:HL-60 Cells; D006801:Humans; D007938:Leukemia / Q000175:diagnosis / Q000235:genetics*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D015514:Oncogene Proteins, Fusion / Q000235:genetics; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D014178:Translocation, Genetic
chemical_list: D015514:Oncogene Proteins, Fusion