A new strategy of discrimination of a point mutation by tandem of short oligonucleotides.

A new strategy based on the use of cooperative tandems of short oligonucleotide derivatives (TSOD) has been proposed to discriminate a "right" DNA target from a target containing a single nucleotide discrepancy. Modification of a DNA target by oligodeoxyribonucleotide reagents was used to characterize their interaction in the perfect and mismatched complexes. It is possible to detect any nucleotide changes in the binding sites of the target with the short oligonucleotide reagent. In the presence of flanking di-3',5'-N-(2-hydroxyethyl)phenazinium derivatives of short oligonucleotides (effectors) the tetranucleotide alkylating reagent modifies DNA target efficiently and site-specifically only in the perfect complex and practically does not modify it in the mismatched complex. It has been shown that TSOD is much more sensitive tool for the detection of a point mutation in DNA as compared to a longer oligonucleotides.

3'-minor groove binder-DNA probes increase sequence specificity at PCR extension temperatures.

DNA probes with conjugated minor groove binder (MGB) groups form extremely stable duplexes with single-stranded DNA targets, allowing shorter probes to be used for hybridization based assays. In this paper, sequence specificity of 3'-MGB probes was explored. In comparison with unmodified DNA, MGB probes had higher melting temperature (T(m)) and increased specificity, especially when a mismatch was in the MGB region of the duplex. To exploit these properties, fluorogenic MGB probes were prepared and investigated in the 5'-nuclease PCR assay (real-time PCR assay, TaqMan assay). A 12mer MGB probe had the same T(m)(65 degrees C) as a no-MGB 27mer probe. The fluorogenic MGB probes were more specific for single base mismatches and fluorescence quenching was more efficient, giving increased sensitivity. A/T rich duplexes were stabilized more than G/C rich duplexes, thereby leveling probe T(m)and simplifying design. In summary, MGB probes were more sequence specific than standard DNA probes, especially for single base mismatches at elevated hybridization temperatures.

[Interaction of derivatives of short oligonucleotides with nucleic acids. VIII. Characteristics of target DNA modification by alkylating oligonucleotide derivatives in tandem complexes]. / Vzaimodeistvie proizvodnykh korotkikh oligonukleotidov s nukleinovymi kislotami. VIII. Osobennosti modifikatsii DNK-misheni v tandemnykh kompleksakh alkiliruiushchimi proizvodnymi oligonukleotidov.

The influence of effectors [octanucleotides and their 3',5'-di-N-(2-hydroxyethyl)phenazinium derivatives] on the modification of a target DNA by alkylating oligonucleotide derivatives forming duplexes of different stability with the target ws studied. It is shown that, being in tandem complexes immediately adjacent to the reactive group of an oligonucleotide reagent possessing a high hybridization capacity, the effector, on the one hand, enhances the stability of the reagent target duplex, and on the other hand, changes the site-specificity of alkylation and decreases the efficiency of the target modification at temperatures that provide a high extent of the target association with the reagent. Conversely, in the case of oligonucleotide reagents forming weak complexes with the target, effectors enhance both the stability of the target.reagent duplex and the extent of the target throughout the temperature range tested. The data indicate that the varying influence of effectors on the target modification by reagents with different hybridization capacities is due to conformational features of the target reagent duplexed regions. Increasing the rigidity of the target.reagent duplex reduces the efficiency of the target modification in tandem complexes.

[Interaction of short oligonucleotide derivatives with nucleic acids. V. Ligation of short oligonucleotides in tandem on a complimentary DNA template]. / Vzaimodeistvie proizvodnykh korotkikh oligonukleotidov s nukleinovymi kislotami. V. Ligirovanie korotkikh oligonukleotidov v tandeme na komplementarnoi matritse DNK.

A tetranucleotide was highly specifically and quantitatively ligated with a pair of flanking octanucleotides carrying both radioactive and nonradioactive reporter groups. The sequence of the ligation of oligonucleotide components in a tandem on a complementary template was studied. The first stage was found to be the enzyme-catalyzed activation of the phosphate group of octanucleotide, a tandem component that possesses a higher hybridization capacity than the tetramer. It is shown that the introduction of terminal reporter groups into octanucleotides does not decrease the efficiency of their tandem ligation.

[Interaction of short oligonucleotides derivatives with nucleic acids. VI. Discrimination of mismatch-containing complexes upon ligation of a short oligonucleotide tandem on DNA template]. / Vzaimodeistvie proizvodnykh korotkikh oligonukleotidov s nukleinovymi kislotami VI. Diskriminatsiia kompleksov, soderzhashchikh mismatch priligirovanii tandema korotkikh oligonukleotidov na DNK-matritse.

The high ligation specificity of a tetranucleotide with a pair of flanking octanucleotides on DNA template by the action of T4 phage DNA ligase is shown. In a tetranucleotide-DNA template complex containing a mismatch, almost no ligation products are formed. The ligation of a tandem octanucleotide-tetranucleotide-octanucleotide makes it possible to identify accurately any single nucleotide substitution in a tetranucleotide binding site.

[Interactions of derivatives of short oligonucleotides with nucleic acids. VII. Effect of conformation changes in the duplex structure on on the specificity and efficacy of modification of target DNA by alkylating oligonucleotide derivatives]. / Vzaimodeistvie proizvodnykh korotkikh oligonukleotidov s nukleinovymi kislotami. VII. Vliianie konformatsionnykh izmenenii struktury dupleksa na napravlennost' i éffektivnost' modifikatsii DNK-misheni alkiliruiushchimi proizvodnymi oligonukleotidov.

The modification of a target DNA by alkylating oligonucleotide derivatives possessing various capacities for complex formation was studied. The binding properties of oligonucleotides were changed either by increasing their length (tetra-, octa-, and dodecamers) or by introducing a point substitution and/or an N-(2-hydroxyethylphenazinium) residue. It was found that conformational changes occurring in the structure of the target.reagent complex upon elevating the reaction temperature affect the efficiency and site-specificity of the alkylation. In the case of complete saturation of the target with the reagent, an increase in the hybridization ability of the reagent reduced the efficiency of the target modification. It was found that the modification by the tetranucleotide reagent (in the presence of an effector adjacent to the 3' end) occurs exclusively at an intracomplex target base. In the case of the dodecamer, which forms a stable, highly cooperative complex with the target, several bases of the target undergo alkylation, and an increase in temperature changes the site-specificity of alkylation. In this process, the redistribution of the target modification sites toward stronger nucleophilic centers enhances alkylation at temperatures near the melting temperature of the target.dodecanucleotide complex despite a decrease in the extent of target association.

Structural studies by high-field NMR spectroscopy of a binary-addressed complementary oligonucleotide system juxtaposing pyrene and perfluoro-azide units.

Recently, a new approach has been proposed to improve the site-specificity and efficiency of the modification of nucleic acid target sequences, the binary system of complementary-addressing nucleic acid sequences. The binary system comprises two oligonucleotides, one modified with a photosensitizing group and the other with a photoreactive group. The sites of chemical modification are arranged to bring the two chemical functions close enough together in space to allow efficient energy transfer from the photo-excited photosensitizer to an arylazide moiety which expels N2 to form a nitrene which subsequently covalently labels the target nucleic acid. Structural analysis performed by high-resolution 2D NMR spectroscopy (400 MHz and 600 MHz) are reported for the model binary system 1:2:3, where 1 is the target 12-mer pdGTATCAGTTTCT, 2 is a photoactivatable fluoroazide derivative dAGAAACp-L-Az and 3 is the photosensitizer derivative Pyr-pdTGATAC (here: Az is the p-azidotetrafluorobenzyl group, Pyr the pyrenyl-1-methylamino group, L a linker group). The assignment of oligonucleotide and modifying group protons was performed using 1H COSY, TOCSY and NOESY experiments. Comprehensive analysis of 1H NOESY spectra of 1:2:3 showed that terminal fragments of the complex [5'p-1T-2G-3A-4T-], [-21A-22T-23A-24C], [-8T-9T-10T-11C-12T] and [13A-14G-15A-15A-17A-18C-] gave a continuous set of intra- and inter-nucleotide interactions, typical of regular double-stranded B-DNA. In contrast, the central region of the complex composed of 5C, 6A, 7G, 19T and 20G nucleotide residues, nearest the Pyr and Az groups, was found to be distorted. Thus some signals from aromatic and/or sugar-ring protons of the above nucleotide residues were extremely broadened or almost absent. Moreover, some intra- and/or inter-nucleotide interactions, typical of the regular DNA duplex, were not detected for the [-5C-6A-7G-] and [-19T-20G-] regions of the tandem system. Instead of that, some cross-peaks of low-intensity between the H2 proton of the Pyr group and 7G(H1'), 7G(H2'/H2"), 7G(H3'), 4T(H2"), 4T(H4') and 4T(H5'/H5") were observed. Additional 1H -1H NOE-interactions between methylene protons of the linker group L and some sugar ring protons of 18C nucleotide residue were detected. A preliminary structural model, constructed using proton-proton distances between Pyr and the DNA and Az-L and DNA obtained from a 1H NOESY experiment at 300 ms mixing time as constraints for the refinement of the structure, displayed significant distortion from B-DNA of the double-stranded helix in the middle of the complex, (-5C-6A-7G, -18C-19T-20G-). The Pyr group was located in what remains of the minor groove near 4T, 5C, 6A and 7G and the centroid of the azide ring less than 9A degrees from the centroid of the ring system of Pyr group.

Modulation of Cm/T, G/A, and G/T triplex stability by conjugate groups in the presence and absence of KCl.

Apparent equilibrium association constants were determined by gel mobility shift analysis for triple strand formation between a duplex target containing a 21 base long A-rich homopurine run and several end-modified C(m)/T (pyrimidine motif; C(m) = 5-methylcytosine), G/A (purine motif), and G/T (purine-pyrimidine motif) triplex-forming oligonucleotides (TFOs). Incubations were carried out for 24 h at 37 degrees C in 20 mM HEPES, pH 7.2, 10 mM MgCl2, and 1 mM spermine. The purine motif triplex was the most stable (Ka = 6.2 x 10(8) M-1) even though the TFO self-associated as a linear duplex. Conjugation of a terminal hexanol or cholesterol group to the G/A-containing TFO reduced triplex stability by 1.6- or 13-fold, whereas an aminohexyl group or intercalating agent (acridine or psoralen) increased triplex stability by 1.3- or 13-fold. These end groups produced similar effects in C(m)/T and G/T triplexes, although the magnitude of the effect sometimes differed. Addition of 140 mM KCl to mimic physiological conditions decreased stability of the G/A triplex by 1900-fold, making it less stable than the C(m)/T triplex. The inhibitory effect of KCl on G/A triplex formation could be partially compensated for by conjugating the TFO to an intercalating agent (30-350-fold stabilization) or by adding the triplex selective intercalator coralyne (1000-fold stabilization). Although the G/T triplex responded similarly to these agents, the stability of the C(m)/T triplex was unaffected by the presence of coralyne and was only enhanced 1.4-2.8-fold when the TFO was linked to an intercalating agent. In physiological buffer supplemented with 40 microM coralyne, the G/A triplex (Ka = 3.0 x 10(8) M-1) was more stable than the C(m)/T and G/T triplexes by factors of 300 and 12, respectively.

Synthesis and hybridization properties of the conjugates of oligonucleotides and stabilization agents--II.

New pyranone derivatives having tri- or pentamethylenamine linker functions were synthesized. These derivatives were covalently attached through the 5'-phosphoramide linkage to heptanucleotide pd(CCAAACA). Complementary complexes of the octanucleotide pd(TGTTTGGC) and above oligonucleotide conjugates were tested for their thermodynamic response. The Tm data and thermodynamic parameters for complex formation have demonstrated the ability of chromone (gamma-pyrone) and coumarin (alpha-pyrone) derivatives to stabilize strongly 7-mer/8-mer complementary complex, most likely through the stacking interaction of the pyran aromatic system with the neighboring nucleotide bases. The effect of chromone (or coumarin) derivatives on the stability of the oligonucleotide complexes (delta delta G at 37 degrees C ranged from -1.0 to -1.7 kcal/mol) was shown to be comparable to the effect of one nucleotide base pair and similar to the effect (delta delta G at 37 degrees C ranged from -1.5 to -2.0 kcal/mol) found for acridineoligonucleotide conjugates served in this study as a reference.

[Selective stabilization of AT-rich DNA duplexes by oligodeoxyribonucleotide conjugates with distamycin analogues]. / Selektivnaya stabilizatsiya AT-bogatykh DNK-dupleksov kon''iugatami oligodezoksiribonuleotidov i analogov distamitsina.

Oligodeoxyribonucleotide conjugates with distamycin analogues containing up to five pyrrolecarboxamide moieties were synthesized. The stability of duplexes formed by these conjugates was shown to depend directly upon the number of pyrrolecarboxamide moieties in the ligand molecule. For the duplexes formed by octaadenylate and octathymidilate conjugates with the distamycin pentapyrrole analogue, stability was demonstrated to be achieved by either one or two ligand molecules; however, duplexes containing two ligand molecules are more stable.

[Nucleic acids interactions with short oligonucleotide derivatives. II. Tandem of short oligonucleotides as highly sensitive system for identification of single base substitutions in target DNA]. / Vzaimodeistvie proizvodnykh korotkikh oligonukleotidov s nykleinovymi kislotami. II. Tandem korotkikh oligonukleotidov--vysokochuvstvitel'naia sistema dlia obnaruzheniia odnobukvennoi zameny v DNK-misheni.

A new approach for modification of target DNAs with tandems of derivatives of short oligonucleotides was suggested that allows highly selective modification of perfect duplexes only. At physiological temperatures, the efficiency of DNA modification by a dodecanucleotide alkylating agent was demonstrated to be the same for both perfect and mismatch-containing duplexes, whereas the tetranucleotide reagent in the presence of two flanking effectors alkylated with high selectivity the target DNA in the perfect duplex only.

[Artificial ribonucleases I. Targeted RNA cleavage by 5'-peptidyloligodeoxyribonucleotides containing arginine and leucine residues]. / Iskustvennye ribonukleazy I. Napravlennoe rasshcheplenie RNK 5'-peptidiloligodezoksiribonukleotidami, soderzhashchimi ostatkami arginina i le'itsina.

The interaction of DNA and RNA with oligodeoxyribonucleotides and their 3'-terminal N-(2-hydroxyethyl)phenazinium derivatives carrying peptide residues with alternating basic and hydrophobic amino acids at the 5'-terminal phosphate was studied. It was found that the introduction of peptide residues (LeuArg)n-Gly-NH2 (n = 2-4) into an oligodeoxyribonucleotide enhances the latter's hybridization ability: each additional LeuArg pair increases the Tm value of the (5')pd(CACACACAAAAAAC).(3')d(TGTGTGTG)p(-LeuArg)n-Gly- NH2 complex by 1.3 degrees C. The reagents did not destort the DNA structure and were capable of site-specific hydrolysis of the phosphodiester bonds of RNA. It was shown that the location of the cleavage sites and the efficacy of the RNA hydrolysis at n = 2 and 4 and at n = 3 strongly differ. The maximum hydrolysis (80%) of tetradecaribonucleotide (5')p(GAUUGAAAAUCCCC) was achieved using peptidyloligodeoxyribonucleotide (3')d(CTAACT)p(LeuArg)4GlyNH2. The possibility of directed cleavage of phosphodiester bonds in tRNAPhe by peptidyloligodeoxyribonucleotides (3')d(CTAACT)p(LeuArg)nGlyNH2 (n = 3 and 4) was shown.

Thermodynamic and spectral properties of DNA miniduplexes with the terminal G x A mispairs and 3' or 5' dangling bases.

The tetradeoxyribonucleotide pAGCG in 1 M NaCl forms duplexes with terminal non-canonical pA x G pairs with stability significantly exceeding that for the duplex (pAGCT)2 and lower than that for the duplex (pCGCG)2. The deoxyriboduplex (pAGCG)2 is considerably stabilized by 3'-Y and slightly by 5'-X dangling bases. Therefore, the stability of duplexes with 3' dangling bases decreases in the order (pAGCGY)2 > (pCGCGA)2 > (pAGCTA)2. The sum of the independent stabilizing effect of the of 5'-pG and 3'-A dangling bases on the (pAGCG)2 core duplex is higher than that of the additional terminal pG x A pair in pG-A-/-G-A tandem of the duplex (pGAGCGA)2.

[Interaction of short nucleotide derivatives with nucleic acids. IV. Modification of DNA by an alkylating tetranucleotide reagents in the presence of effectors in perfect and imperfect complexes]. / Vzaimodeistvie proizvodnykh korotkikh oligonukleotidov s nukleinovymi kislotami. IV. Modifikatsiia DNK alkiliruiushchim proizvodnym tetranucleotida v prisutstvii éffektorov v sostave pravil'nykh i nepravil'nykh kompleksov.

It was demonstrated that any mismatches in a complex formed by an ssDNA target and a tetranucleotide at 25 or 37 degrees C can be discriminated by alkylating the DNA with a tetranucleotide carrying a 4-[N-methyl-N-(2-chloroethyl)]aminobenzylethylamine residue at the 5'-terminal phosphate in the presence of a pair of flanking effectors, octanucleotide di-N-(2-hydroxyethyl)-phenazinium derivatives. The discrimination factor (ratio of the extent of the target modification in the perfect and mismatch-containing complexes) for a single mismatch in the tetranucleotide binding site at 25 degrees C varied between 4 and 500 depending on the type of mismatch and its location in the complex and exceeded 400 at 37 degrees C for all the investigated mismatches. The DNA target modification by the alkylating derivative of the 3'-estrone ester of tetranucleotide pCAGX (mean = C, T, A or G) was selective in the presence of a pair of hydrophobic effectors, octanucleotide 5'-cholesteryl-3'-phenazinium derivatives. The discrimination factors for 3'-terminal mismatches T.G, A.G, and G.G were 1,8,400, and 400, respectively.

[Interaction of derivatives of short oligonucleotides with nucleic acids. I. Effect of various types of effectors on alkylation of DNA-targets]. / Vzaimodeistvie proizvodnykh korotkikh oligonukleotidov s nukleinovymi kislotami. I. Vliianie razlichnykh tipov éffektorov na alkilirovanie DNK-mishenei.

It was shown that the tandem of the derivatives of short oligonucleotides efficiently and site specifically interacts with target 20 base deoxyribonucleotide (M). It was demonstrated that the very low hybridization ability of tetranucleotide (D) and its 3'-cholesterol and 3'-estrone esters (D-ChS and D-EsS, respectively) increases significantly in the presence of the effectors: octanucleotides (E1 and E2), and their 5',3'-diphenazinium (Phn-E1-Phn and Phn-E2-Phn) and 5'-cholesteryl-3'-phenazinium (ChS-E1-Phn and ChS-E2-Phn) derivatives, which flank them on the target strand. The influence of the effectors on the interaction of the target M with tetranucleotide D or its alkylating derivatives (RCl-D) increases in a series E1 + E2 < ChS-E1-Phn + ChS-E2-Phn < Phn-E1-Phn + Phn-E2-Phn. For the steroid derivatives, D-ChS and D-EsS, and the reagents based on them (RCl-D-ChS and RCl-D-EsS), this series is E1 + E2 < Phn-E1-Phn + Phn-E2-Phn < ChS-E1-Phn + ChS-E2-Phn. The modification level of the target M with derivatives RCl-D-EsS in the presence of ChS-E1-Phn and ChS-E2-Phn reaches 40% even at 37 degrees C under conditions close to physiological. The possibility of using 5'-cholesteryl-3'-phenazinium-containing oligonucleotides as effectors of the interaction of target DNA with the derivatives of short oligonucleotides was demonstrated.

[Effect of the intercalating dyes ethidium and phenazine, covalently jointed to the 5'- or 3'-end of the pentanucleotide d(pGAAAG), on the thermodynamics of complementary and cooperative interaction]. / Vliianie interkaliruiushchikh krasitelei étidiia i fenaziniia, kovalentno prisoedinennykh k 5'- ili 3'-kontsu pentanukleotida d(pGAAAG), na termodinamiku komplementarnogo i kooperativnogo vzaimodeistviia.

Thermal stability was studied and thermodynamic parameters of complex formation were calculated for pentanucleotide complexes [formula: see text] and corresponding complexes of pentanucleotide derivatives carrying at their 5'- or 3'-ends covalently bound residues of intercalating dyes: N-(2-hydroxyethyl)-phenazine (Phn) or 2-N-(3-aminopropionyl)-ethidium. Pentanucleotide derivatives were shown to form more stable complementary complexes. The best stabilizing effect was observed when the dye was oriented towards the long single-stranded fragment of tetradecanucleotide, melting temperature of the complexes being by 22.5 (Phn) and 31.2 degrees C (Etd) greater than that of unmodified complexes in the case of 5'-derivatives and by 21.6 (Phn) and 27.2 degrees C (Etd) for 3'-derivatives. Cooperativity constant of pentanucleotide derivatives in "tandem" complexes IV-VII was higher than that of unmodified pentanucleotide. For complex IV at 37 degrees C the constant values were 33 (unmodified), 35 (5'Phn), 57 (3'Phn), 190 (5'Etd), 100 (3'Etd). With n = 3 in complex VII cooperativity constants approached 1 in any case.

Ethidium and azidoethidium oligonucleotide derivatives: synthesis, complementary complex formation and sequence-specific photomodification of the single-stranded and double-stranded target oligo- and polynucleotides.

Oligodeoxyribonucleotide derivatives containing ethidium or azidoethidium residues attached to 3' and/or 5' end were prepared. These derivatives formed tight specific complexes with complementary oligodeoxyribonucleotides where each attached ethidium residue led to an increase of complex Tm by 20-30 degrees C. Tandem complexes of two oligodeoxyribonucleotides containing ethidium residues with an oligodeoxyribonucleotide having two adjacent complementary sequences for these oligonucleotides were investigated. Photoinduced reactions of a number of ethidium and azidoethidium oligodeoxyribonucleotide derivatives with target complementary single-stranded and double-stranded oligo- and polydeoxyribonucleotides were investigated. The irradiation led to direct photocleavage of the target oligo- or polynucleotide, to formation of hidden (piperidine cleavable) modifications of the target and to formation of covalent adducts between ethidium oligodeoxyribonucleotide derivative and the target. In a number of experiments, azidoethidium dyes were demonstrated to be considerably stronger photosensitizers than ethidium ones. Depending on the nature of the target (single- or double-stranded DNA) and on the irradiation conditions, the total damages to the target oligo- or polydeoxyribonucleotides ranged from 10-70% (for ethidium dyes) to 30-80% (for azidoethidium dyes).

Synthesis of specific diastereomers of a DNA methylphosphonate heptamer, d(CpCpApApApCpA), and stability of base pairing with the normal DNA octamer d(TPGPTPTPTPGPGPC).

DNA methylphosphonates are candidate derivatives for use in antisense DNA therapy. Their efficacy is limited by weak hybridization. One hypothesis to explain this phenomenon holds that one configuration of the chiral methylphosphonate linkage, Rp, permits stronger base pairing than the other configuration, Sp. To test this hypothesis, four specific pairs of Rp and Sp diastereomers of the DNA methylphosphonate heptamer d(CpCpApApApCpA) were prepared by block coupling of different combinations of individual diastereomers of d(CpCpApA) and d(ApCpA). Each pair of the diastereomers of the heptamer was separated into individual diastereomes using affinity chromatography on a Lichrosorb-NH2 silica column with a covalently attached complementary normal DNA octamer, d(pTpGpTpTpTpGpGpC). The stabilities of complementary complexes of phosphodiester d(TpGpTpTpTpGpGpC) with 8 individual diastereomers of methylphosphonate d(CpCpApApApCpA) were studied by measuring their melting temperatures (Tm). A direct correlation of Tm values with the number of Rp methylphosphonate centers in the heptamer was found: the more Rp centers, the higher the stability of the complex. Tm values for the diastereomers with 6 all-Rp or all-Sp methylphosphonate centers were found to be 30.5 degrees and 12.5 degrees C, respectively, in 100 mM NaCl, 10 mM Na2HPO4, 1 mM EDTA, pH 7.0 with 15 microM of each oligomer. On the average, each substitution of one Rp-center to an Sp-center in the heptamer decreased the Tm by 3 degrees C. Under the same conditions, the Tm of the normal DNA heptamer with its complement was 21 degrees C. These results are consistent with the model that all-Rp methylphosphonate DNAs hybridize much more tightly to complementary normal DNA than do racemic methylphosphonate DNAs, and may therefore exhibit greater potency as antisense inhibitors.

[Complementary-addressed photomodification of DNA-targets by arylazide and perfluoroarylazide oligonucleotide derivatives. IV. Photomodification of ss- and ds-DNA-fragments]. / Komplementarno-adresovannaia fotomodifikatsiia DNK-mishenei arilazidnymi i perftorarilazidnymi proizvodnymi oligonukleotidov. IV. Fotomodifikatsiia ss- i ds-DNK-fragmentov.

A highly efficient sequence-specific photomodification of single stranded (ss) and double stranded (ds) DNA fragments was carried out with hexadecathymidilate derivative, R-p(T)16(R--p-azidotetrafluorobenzamide) and 27-meric DNA fragments as a targets. [formula: see text] The main points of the modification were G7 and G24 for the ss target and G7 and G22 of purine- and pyrimidine-rich strands, respectively, for the ds DNA fragment. The photomodification extent was 60-77% for ss DNA and 10-53% for ds DNA depending on the reaction conditions: it increased in a buffer with a high ionic strength (1.0 M) and at a low temperature (4 degrees C) when the triplexes are more stable.