Russian Journal of Bioorganic Chemistry, Vol. 31, No. 2, 2005, p.170

A Method for the Preparation of Normalized cDNA Libraries Enriched with Full-Length Sequences

P. A. Zhulidov 1 , E. A. Bogdanova 1 , A. S. Shcheglov 1 , I. A. Shagina 2 , L. L. Wagner 1, 2 ,G. L. Khazpekov 3 , V. V. Kozhemyako 4 , S. A. Lukyanov 1 ,* and D. A. Shagin 1

1 Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry , Russian Academy of Sciences ,ul. Miklukho-Maklaya 16/10, Moscow, 117997 Russia, 2 ZAO Evrogen, Moscow, Russia, 3 Cardiological Research Center, Russian Academy of Medical Sciences, Moscow, Russia, 4 Pacific Institute of Bioorganic Chemistry, Far East Division, Russian Academy of Sciences, pr. Stoletiya Vladivostoka 159, Vladivostok, 690022 Russia

Abstract: We developed a new method for the preparation of normalized cDNA libraries enriched with full-length sequences. It is based on the properties of the recently characterized duplex-specific nuclease from the hepatopancreas of the Kamchatka crab. The duplex-specific nuclease is thermostable, effectively cleaves double-stranded DNA, and is inactive toward single-stranded DNA (Shagin et al., Genome Res., 2002, vol. 12, pp.1935–1942). Our method enables the normalization of cDNA samples enriched with full-length sequences without use of laborious and ineffective stages of physical separation. The efficiency of the method was demonstrated in model experiments using cDNA samples from several human tissues.

Key words: cDNA normalization, duplex-specific nuclease, normalized cDNA libraries