The effect of interior position on the free energy increment for a single LNA substitution for a 2 0 -O-methyl RNA was studied for the duplex 5 0 A M C M U M A M C M C M A M /3 0 r(UGAUGGU). As summarized in Table 1 , a single interior LNA substitution makes duplex stability more favorable by 1.2-1.7 kcal/mol at 37 C, with an average of 1.4 kcal/mol. This corresponds to roughly a 10-fold increase in binding constant. Thus, interior and 3 0 terminal LNA substitutions usually improve binding more than 5 0 terminal LNA substitutions. Table 1 . For 13 of 16 sequences, the LNA substitution makes duplex stability more favorable by 1.0-1.5 kcal/mol at 37 C, with an average enhancement of 1.3 kcal/mol. The enhancement for the other three sequences averages 2.1 kcal/mol at 37 C.