Selective alkylation of poly(A) tracts of RNA inside the cell with the derivative of ethyl ester of oligothymidilate bearing 2‐Chloroethylamino group

The method of the complementary addressed modification was proposed [I] and realised [2-41 by N. I. Grineva et al. to perform the selective labeling of single-stranded nucleic acids at the predetermined points. The methods were elaborated for the binding of the reactive 2-chloroethylamino group to either 3’or 5’-end of oligonucleotides directing the reagent to the chosen region of nucleic acid [5,6]. The selective mutagenesis of T7 DNA has been described, based on the mRNA derivatives with 2-chloroethylamino groups attached to the guanine residues [7]. A similar idea was presented in [8]. To employ complementary addressed modification in situ it is necessary to overcome at least two difficulties : (i) The low ability of the hydrophilic oligonucleotide or polynucleotide derivatives to cross the outer cell membrane; (ii) The sensitivity of the oligonucleotide moiety of the reagents of the above type to intracellular nucleases. Esterification of the.internucleotide phosphates with ethanol does not result in the loss of the ability of oligonucleotides to undergo Watson-Crick basepairing [9,10]. At the same time these nonzcharged oligonucleotide analogs are sufficiently hydrophobic to penetrate through the cell membrane and exhibit rather high stability towards digestion by nucleases [ill. Here, we describe the alkylating pentanucleotide derivative (CsHSNH)*p(dTpEt)4U with completely esterified internucleotide phosphates and with 2’,3’0-4-(N-2-chloroethyl-N-methylamino)-benzylidene residue attached to the 3’-end of the molecule.