For four cases, the effect of a mismatch with an LNA nucleotide was compared with that for the equivalent 2 0 -O-methyl nucleotide. In each case, the mismatch penalty for the LNA was less than that for 2 0 -O-methyl RNA. However, for an A M -G mismatch flanked by LNAs in the context 5 0 A L C M U L A M C L C M A L /3 0 r(UGAGGGU), the LNAs enhanced the mismatch penalty by $1 kcal/mol relative to a completely 2 0 -O-methyl RNA strand. Thus, oligonucleotides containing LNA may discriminate best against mismatches flanked by LNAs.