The evaluation of a new method for detecting fusion genes, using MLL gene rearrangement as an example

• Main Authors: Yin-JuLin, 林殷如
• Other Authors: Chung-Liang Ho
• Format: Others
• Language: zh-TW
• Published: 2010
• Online Access: http://ndltd.ncl.edu.tw/handle/77350949150454477274

碩士 === 國立成功大學 === 分子醫學研究所 === 99 === Fusion genes, resulting from chromosome translocations, are the most prevalent form of somatic changes in the cancer genome. Intriguingly, ＞80% of all known fusion genes are attributed to leukemias, lymphomas, but bone and soft tissue sarcomas that account for only 10% of all human cancers. In contrast, common epithelial cancers, which account for 80% of cancer-related deaths, can only be attributed to 10% of known recurrent fusion genes. Furthermore, if translocation-mediated fusion genes encode activated enzymes with direct oncogenic potential, targeting such enzymes could provide a feasible approach to treat individuals harboring the corresponding fusion genes. Therefore, a high-throughput method is needed not only for differential diagnosis but also for discovering drugable fusion genes in solid tumors. RACE (rapid amplification of cDNA ends) is a widely used technique for finding fusion genes but its chief defect is the demand of a great deal of sequecing which may result from low fusion/wild-type transcript ratios. We propose to use SAGE (serial analysis of gene expression) after RACE to increase the throughput. When sequencing same amount of clones, SAGE can give 20 to 30 times more information than RACE alone. Therefore, it would be possible to pool the cases into groups and screened at once. This strategy is designated SAAT (serial analysis of amplified transcripts). In order to assess the performance of SAAT we use MLL (Mixed Lineage Leukaemia) translocations as an example. MLL translocations occur in up to 70% of infant Acute lymphoblastic leukemia (ALL), and in about 3% of acute myeloid leukemia (AML), and it is already known can be recombined with 64 different partner genes, posing a challenge for routine molecular diagnosis. In this research we based on the intergraty of the basic reaserch of MLL gene, sucessfully established a RACE system specific to MLL gene.Then we mixed normal leukocyte and MV4-11 that have MLL-AF4 translocation at different ratio to test the correctness and the detection efficiency of SAAT.In the future, we well use SAAT to screen a pool of samples from carcinoma patients at a time, for the purpose of discovering drugable fusion genes in solid tumors.