Functionalized Nanocomposite Coating of a Glass Surface for Oligonucleotide Immobilization

V. V. Annenkova, A. S. Levinab, E. N. Danilovtsevaa, E. A. Filinaa, E. A. Mikhalevab, and V. F. Zarytovab,#

# Phone: +7 (383) 335-6224; fax: +7 (383) 333-3677; e-mail:
a Limnological Institute, Siberian Division, Russian Academy of Sciences, ul. Lermontova 281, Irkutsk, 664033 Russia
b Institute of Chemical Biology and Fundamental Medicine, Siberian Division, Russian Academy of Sciences, pr. Lavrent’eva 8, Novosibirsk, 630090 Russia

Received June 22, 2005; in final form, August 8, 2005

Abstract: A new type of coating for manufacturing DNA chips was constructed on the basis of an organic–inorganic nanocomposite based on the polyvinylbutyral–tetraethoxysilane copolymer. The organosilicon composite was functionalized by introduction of ethanolamine vinyl ether copolymers, which contain amino groups and anchor vinyloxide units capable of reacting with silanol groups of the nanocomposite. The resulting coatings form a film on glass slides with a high surface density of amino groups (up to 700 groups/nm2) suitable for three-dimensional immobilization of oligonucleotides. The use of bifunctional reagents (e.g., phenylene diisothiocyanate) for the attachment of oligonucleotides bearing amino linkers to the amino-containing surface provides an immobilization density of 0.5--1.6 pmol/mm2. Immobilization with a higher density (10--12 pmol/mm2) was achieved for attachment to amino-containing glass slides upon the use of oligonucleotides containing a selectively activated terminal phosphate group. The activation of oligonucleotides was carried out with the triphenylphosphine–dithiodipyridine pair in the presence of dimethylaminopyridine N-oxide. The resulting DNA chips were shown to be useful in principle for DNA detection.

Key words: DNA chips, immobilization, nanocomposite, oligonucleotides, polysiloxane, polyvinylbutyral

Russian Journal of Bioorganic Chemistry 2006, 32 (5):460-467