Optimal design of parallel triplex forming oligonucleotides containing Twisted Intercalating Nucleic Acids--TINA

TY - JOUR

T1 - Optimal design of parallel triplex forming oligonucleotides containing Twisted Intercalating Nucleic Acids--TINA

AU - Schneider, Uffe V

AU - Mikkelsen, Nikolaj D

AU - Jøhnk, Nina

AU - Okkels, Limei M

AU - Westh, Henrik

AU - Lisby, Gorm

PY - 2010/3/24

Y1 - 2010/3/24

N2 - Twisted intercalating nucleic acid (TINA) is a novel intercalator and stabilizer of Hoogsteen type parallel triplex formations (PT). Specific design rules for position of TINA in triplex forming oligonucleotides (TFOs) have not previously been presented. We describe a complete collection of easy and robust design rules based upon more than 2500 melting points (Tm) determined by FRET. To increase the sensitivity of PT, multiple TINAs should be placed with at least 3 nt in-between or preferable one TINA for each half helixturn and/or whole helixturn. We find that ΔTm of base mismatches on PT is re- markably high (between 7.4 and 15.2oC) compared to antiparallel duplexes (between 3.8 and 9.4oC). The specificity of PT by "Tm increases when shorter TFOs and higher pH are chosen. To increase ΔTms, base mismatches should be placed in the center of the TFO and when feasible, A, C or T to G base mismatches should be avoided. Base mismatches can be neutralized by intercalation of a TINA on each side of the base mismatch and masked by a TINA intercalating direct 3' (preferable) or 5' of it. We predict that TINA stabilized PT will improve the sensitivity and specificity of DNA based clinical diagnostic assays.

AB - Twisted intercalating nucleic acid (TINA) is a novel intercalator and stabilizer of Hoogsteen type parallel triplex formations (PT). Specific design rules for position of TINA in triplex forming oligonucleotides (TFOs) have not previously been presented. We describe a complete collection of easy and robust design rules based upon more than 2500 melting points (Tm) determined by FRET. To increase the sensitivity of PT, multiple TINAs should be placed with at least 3 nt in-between or preferable one TINA for each half helixturn and/or whole helixturn. We find that ΔTm of base mismatches on PT is re- markably high (between 7.4 and 15.2oC) compared to antiparallel duplexes (between 3.8 and 9.4oC). The specificity of PT by "Tm increases when shorter TFOs and higher pH are chosen. To increase ΔTms, base mismatches should be placed in the center of the TFO and when feasible, A, C or T to G base mismatches should be avoided. Base mismatches can be neutralized by intercalation of a TINA on each side of the base mismatch and masked by a TINA intercalating direct 3' (preferable) or 5' of it. We predict that TINA stabilized PT will improve the sensitivity and specificity of DNA based clinical diagnostic assays.

M3 - Journal article

SN - 0305-1048

VL - 38

SP - 4394

EP - 4403

JO - Nucleic Acids Research

JF - Nucleic Acids Research

IS - 13

ER -