High throughput detection of microsatellite instability (MSI) in sporadic colorectal cancer by MSI COPPER denaturing high performance liquid chromatography.

摘要

Hereditary nonpolyposis colorectal cancer (HNPCC) is the most common hereditary colorectal cancer (CRC) syndrome, which is caused by germline mutations in the mismatch repair (MMR) genes hMLHl and hMSH2 (http://www.insight-group.org/, accessed June 2008). Persons inheriting this trait are particularly at risk of developing cancer of the colon, rectum, endometrium, small bowel and urinary tract. In addition, microsatellite instability (MSI), which is characterised by differences in the length of repeated sequences between normal and tumour DNA from the same individual, is caused by MMR replication errors. In the research and clinical laboratory setting of HNPCC, analysis for MSI status is applied as a prescreening test prior to mutation analysis in hMLHl and hMSH2 genes. Recently, several electrophoretic methods have been used for determination of MSI status.3'4 While these methods can provide simultaneous amplification and detection of the MSI markers, major drawbacks include: (1) the need for fluorescently-labelled primers which increase the cost of analysis, or the use of radiation-labelled primers; (2) due to the different annealing temperatures of traditional primer sets, amplicons for MSI markers cannot be generated under the same PCR thermal cycler condition which drastically increases sample preparation time; (3) optimal conditions for MSI analysis of each of the amplified products are not provided through instrumental software of the WAVE system (Transgenomic, USA), which increase both the cost of analysis and the technician's health risk.