CHAPTER 10 CURRENT ATTEMPTS TO IMPROVE THE SPECIFICITY OF NUCLEIC ACIDS HYBRIDIZATION

Abstract: Being very useful and informative, many techniques based on nucleic acids hybridization suffer from the cross-annealing of repetitive DNA, presenting in reassociating samples. This “wrong ” annealing causes “nonspecific ” hybridization of nonorthologous DNA fragments, thus producing chimeric sequences and at the final stage significantly hampering the analysis of the resulting cDNA or genomic libraries. Such chimeras may constitute up to 40–60 % of DNA libraries. Importantly, the number of chimerical clones positively correlates with the com-plexity of hybridizing genomic or cDNA mixtures. The hybridization specificity is a crucial factor determining both the fidelity the efficiency of all hybridization-based analytical techniques. In this chapter, I review the current attempts to increase the specificity of hybridization at both stages: during nucleic acids reasso-ciation and at the stage of selection of proper hybrids. To this end, approaches based on chemical modifications, improving hybridization kinetics, and improving selection of perfectly matched duplexes, have been developed