Minimising the secondary structure of DNA targets by incorporation of a modified deoxynucleoside: implications for nucleic acid analysis by hybridisation
Open Access
- 15 October 2000
- journal article
- research article
- Published by Oxford University Press (OUP) in Nucleic Acids Research
- Vol. 28 (20) , 3904-3909
- https://doi.org/10.1093/nar/28.20.3904
Abstract
Some regions of nucleic acid targets are not accessible to heteroduplex formation with complementary oligonucleotide probes because they are involved in secondary structure through intramolecular Watson–Crick pairing. The secondary conformation of the target may be destabilised to assist its interaction with oligonucleotide probes. To achieve this, we modified a DNA target, which has self-complementary sequence able to form a hairpin loop, by replacing dC with N4-ethyldeoxycytidine (d4EtC), which hybridises specifically with natural dG to give a G:4EtC base pair with reduced stability compared to the natural G:C base pair. Substitution by d4EtC greatly reduced formation of the target secondary structure. The lower level of secondary structure allowed hybridisation with complementary probes made with natural bases. We confirmed that hybridisation could be further enhanced by modifying the probes with intercalating groups which stabilise the duplex.Keywords
This publication has 22 references indexed in Scilit:
- Resequencing and mutational analysis using oligonucleotide microarraysNature Genetics, 1999
- Arrays of complementary oligonucleotides for analysing the hybridisation behaviour of nucleic acidsNucleic Acids Research, 1994
- Studies of oligonucleotide interactions by hybridisation to arrays: the influence of dangling ends on duplex yieldNucleic Acids Research, 1994
- Elimination of band compression in sequencing gels by the use of N4-methyl-2′-deoxycytidine 5′-triphosphateNucleic Acids Research, 1993
- 7-Deaza-2′-deoxyadenosine and 3-deaza-2′-deoxyadenosine replacing dA within d(A6)-tracts: differential bending at 3′-and 5′-junctions of d(A6) d(T6) and B-DNANucleic Acids Research, 1992
- Specific inhibition of mRNA translation by complementary oligonucleotides covalently linked to intercalating agents.Proceedings of the National Academy of Sciences, 1986
- Nucleic acid-binding molecules with high affinity and base sequence specificity: intercalating agents covalently linked to oligodeoxynucleotides.Proceedings of the National Academy of Sciences, 1984
- Poly(7-deazaguanylic acid), the homopolynucleotide of the parent nucleoside of queuosineBiochemistry, 1982
- Modified porynudeotides. VI. Properties of a synthetic DNA containing the anti-herpes agent (E)-5-(2-bromovinyI)-2′-deoxyuridineNucleic Acids Research, 1982
- Recognition sequences of repressor and polymerase in the operators of bacteriophage lambdaCell, 1975