Table 3.

Design rules for placement of TINA in PT

For maximum increase in ΔTm for the TFO (increase sensitivity)

(1) Place the TINAs in the TFO and internal TINAs as a bulge (ref. no. 1)

(2) Place a TINA at each end of the oligonucleotide

(3) Always place TINAs with three or more nucleotides in-between

(4) Place a TINA for each half helixtum and/or whole helixtum

(5) For pH > 6.0 use maximum number of TINAs following rules 3 and 4

(6) For oligonucleotides shorter than 9 nts use rule 2

To increase ΔTm for the TFO in parallel triplex (increase specificity)

(1) Position target base mismatches in the centre of the TFO

(2) Avoid A, C or T to G target base mismatches when feasible

(3) Choose a shorter TFO or higher pH to increase ΔTm of a target base mismatch

(4) Place TINAs several nucleotides from the target base mismatch

To decrease ΔTm for the TFO in parallel triplex (neutralize effect of base mismatch)

(1) Place a TINA direct 3′ of a target base mismatch to reduce ΔTm

(2) Place two TINA's direct 5′ and 3′ of a target base mismatch to neutralize ΔTm