Use of DNA chromatography for sequence analysis

S.P. Yip,¹ K.H. Leung,¹ P.M. Ngan,¹ W.C. Tang^{1, 2} and M.K.H. Yap²

¹Department of Health Technology and Informatics, and
²School of Optometry, The Hong Kong Polytechnic University.

Analysis of DNA sequence variations, either polymorphisms or disease-causing mutations, is crucial to biomedical science research and to molecular testing for the diagnosis of diseases. DNA chromatography differentially separates DNA molecules, double- or single-stranded, on the basis of their interaction through amphiphilic ions with the non-polar stationary phase and the polar mobile phase. Under non-denaturing conditions, DNA chromatography separates DNA molecules according to their size. Under partially denaturing conditions, DNA chromatography can differentially separate homoduplexes and heteroduplexes carrying mismatches under suitable column temperature, which varies according to the sequence of the DNA fragments being analysed. As such, it has now become a very powerful tool for screening and identifying unknown sequence variations. Under completely denaturing conditions, DNA chromatography separates short single-stranded DNA fragments according to both the length and base composition of the fragments. Thus, coupled with primer extension reaction, it allows rapid and reliable genotyping of known sequence variations. Multiplexing of primer extension reactions can increase the throughput and is particularly useful in molecular diagnostics. On the other hand, DNA chromatography can also be used in conjunction with DNA pooling. Under partially denaturing conditions, DNA pooling can speed up the process of screening sequence variations. Under completely denaturing conditions, DNA pooling allows accurate estimation of relative allele frequencies of single nucleotide polymorphisms analysed by primer extension coupled with DHPLC. This approach is extremely useful in mapping genes involved in complex diseases using genetic association studies.

This article serves to give a brief account of the principle underlying DNA chromatography and the use of this technology for DNA analysis in both research and diagnostic settings. References have been purposely kept to a minimum in the main text. Interested readers can refer to two recent excellent reviews, on by Xiao and Oefner¹ and another by Gjerde and co-workers².

Keywords: DNA chromatography, denaturing high performance liquid chromatography, polymorphisms, mutations.

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