Yuri P. Lysov, Fedor N. Gnuchev, Andrei A. Mironov, Alexey A. Chernyi, and Andrei D. Mirzabekov
Argonne National Laboratory (U.S.A.) and Engelhardt Institute of Molecular Biology (Moscow)-Joint Human Genome Program.
DNA sequencing by hybridization on oligonucleotide microchips (SHOM) allows the determination of a spectrum of overlapping oligonucleotides constituting a DNA fragment. The oligomers that hybridize to form perfect duplexes with an array of immobilized oligonucleotides and, as a result, reconstitution of the nucleotide sequence of the fragment is possible. In longer DNA fragments, unambiguous reconstitution of a DNA sequence is often impeded by the presence of repetitive regions and simple sequence repeats. Here it is demonstrated that SHOM, supplemented by measurement of the distance between certain sites within the analyzed DNA (for example, restriction sites or priming sites for PCR), enables sequencing of much longer DNA fragments containing repeats of varying complexity. Results of computer simulations on sequences from the EMBL database in the context of a model experiment including contiguous stacking hybridization are presented. These results show expected efficiency of the method as function of the sequence origin and are intended to be used as guidelines for real sequencing experiment planning. Sequences up to 10,000 bp long are reconstructed efficiently.
*Work supported in part by the U.S. Department of Energy, Office of Health and Environmental Research, under Contract No. W-31-109-ENG-38 and Russian Human Genome Program.
The submitted manuscript has been authorized by a contractor of the U.S. Government under contract No. W-31-109-ENG-38. Accordingly, the U.S. Government retains a nonexclusive, royalty-free license to publish or reproduce the published form of this contribution, or allow others to do so, for U.S. Government purposes.