[Kinetic analysis for subtractive hybridization of transcripts]

A theoretical analysis of kinetics of two subtractive cDNA hybridizations has been carried out: hybridisation of an excess of denatured double-stranded cDNA (driver) with denatured double-stranded cDNA (tracer) containing some sequences (targets) missing from the driver, and hybridisation of denatured double-stranded tracer with denatured double-stranded driver unable to be renatured. Our calculations show that the first strategy requires several rounds of hybridisation for sufficient enrichment of tracer with target sequences of the low-abundant mRNA: in the first round the renatured fraction is removed, and in subsequent rounds the remaining single-stranded fraction cDNA is used and renatured fraction is collected. The second strategy makes it possible to sufficiently enrich non-reassociated fraction of tracer with minor fractions of target sequences in a single round of subtractive hybridisation.