LNA oligonucleotides exhibit unprecedented thermal stability when hybridized to a complementary DNA or RNA strand. For each incorporated LNA monomer, the melting temperature (Tm
) of the duplex increases by 2–8°C (see figure Replace DNA with LNA for higher melting temperature
). In addition, LNA oligonucleotides can be made shorter than traditional DNA or RNA oligonucleotides and still retain a high Tm
. This is important when the oligonucleotide is used to detect small or highly similar targets.
Since LNA oligonucleotides typically consist of a mixture of LNA and DNA or RNA, it is possible to optimize the sensitivity and specificity by varying the LNA content of the oligonucleotide. Incorporation of LNA into oligonucleotides has been shown to improve sensitivity and specificity for many hybridization-based technologies including PCR, microarrays and in situ hybridization (ISH).
normalization enables robust detection, regardless of GC content. The Tm
of a nucleotide duplex can be controlled by varying the LNA content. This feature can be used to normalize the Tm
across a population of short sequences with varying GC content. For AT-rich nucleotides, which give low melting temperatures, more LNA is incorporated into the LNA oligonucleotide to raise the Tm
of the duplex. This enables the design of LNA oligonucleotides with a narrow Tm
range, which is beneficial in many research applications such as microarrays, PCR and other applications in which sensitive and specific binding to many different targets must occur under the same conditions simultaneously. The power of Tm
normalization is demonstrated by the comparison of DNA and LNA probes for detection of miRNA targets with a range of CG content (see figure The power of Tm normalization