Locked nucleic acid
Encyclopedia
A locked nucleic acid (LNA), often referred to as inaccessible RNA, is a modified RNA
nucleotide
. The ribose moiety of an LNA nucleotide is modified with an extra bridge connecting the 2' oxygen and 4' carbon. The bridge "locks" the ribose in the 3'-endo (North) conformation, which is often found in the A-form duplexes. LNA nucleotides can be mixed with DNA or RNA residues in the oligonucleotide whenever desired. Such oligomers are synthesized chemically
and are commercially available. The locked ribose conformation enhances base stacking and backbone pre-organization. This significantly increases the hybridization properties (melting temperature
) of oligonucleotide
s.
LNA was independently synthesized
by the group of Jesper Wengel in 1998, soon after the first synthesis by the group of Takeshi Imanishi in 1997. The exclusive rights to the LNA technology were secured in 1997 by Exiqon A/S, a Danish biotech company.
LNA nucleotides are used to increase the sensitivity and specificity of expression in DNA microarray
s, FISH probes, real-time PCR probes and other molecular biology techniques based on oligonucleotides. For the in situ detection of miRNA
the use of LNA is currently (2005) the only efficient method. A triplet of LNA nucleotides surrounding a single-base mismatch site maximizes LNA probe specificity unless the probe contains the guanine base of G-T mismatch.
Using LNA based oligonucleotides therapeutically is an emerging field in biotechnology
. The Danish pharmaceutical company Santaris Pharma a/s
owns the sole rights to therapeutic uses of LNA technology, and is now developing a new, LNA based, hepatitis C
drug called miravirsen, targeting miR-122
, which is in Phase II clinical testing as of late 2010.
Benefits of the LNA technology
Some of the benefits of using LNA include:
Ideal for the detection of short RNA and DNA targets
Increases the thermal stability of duplexes
Capable of single nucleotide discrimination
Resistant to exo- and endonucleases resulting in high stability in vivo and in vitro applications
Increased target specificity
Facilitates Tm normalization
Strand invasion properties enables detection of “hard to access” samples
Compatible with standard enzymatic processes
Applications of the LNA technology
Some proven applications of LNA include:
Allele-specific PCR: allows for the design of shorter primers, without compromising binding specificity
Microarray gene expression profiling: provides increased sensitivity and selectivity with smaller amounts of substrates
Small RNA research
SNP genotyping
mRNA antisense oligonucleotides
RNAi
DNAzymes
Fluorescence Polarization probes
Molecular Beacons
Gene repair/exon skipping
Splice variant detection
Comparative genome hybridization (GCH)
Other therapeutic and diagnostic applications of LNA technology are in development.
RNA
Ribonucleic acid , or RNA, is one of the three major macromolecules that are essential for all known forms of life....
nucleotide
Nucleotide
Nucleotides are molecules that, when joined together, make up the structural units of RNA and DNA. In addition, nucleotides participate in cellular signaling , and are incorporated into important cofactors of enzymatic reactions...
. The ribose moiety of an LNA nucleotide is modified with an extra bridge connecting the 2' oxygen and 4' carbon. The bridge "locks" the ribose in the 3'-endo (North) conformation, which is often found in the A-form duplexes. LNA nucleotides can be mixed with DNA or RNA residues in the oligonucleotide whenever desired. Such oligomers are synthesized chemically
Oligonucleotide synthesis
Oligonucleotide synthesis is the chemical synthesis of relatively short fragments of nucleic acids with defined chemical structure . The technique is extremely useful in current laboratory practice because it provides a rapid and inexpensive access to custom-made oligonucleotides of the desired...
and are commercially available. The locked ribose conformation enhances base stacking and backbone pre-organization. This significantly increases the hybridization properties (melting temperature
Melting temperature
Melting temperature may refer to:* Melting point, the temperature at which a substance changes from solid to liquid state.* DNA melting temperature, the temperature at which a DNA double helix dissociates into single strands....
) of oligonucleotide
Oligonucleotide
An oligonucleotide is a short nucleic acid polymer, typically with fifty or fewer bases. Although they can be formed by bond cleavage of longer segments, they are now more commonly synthesized, in a sequence-specific manner, from individual nucleoside phosphoramidites...
s.
LNA was independently synthesized
Oligonucleotide synthesis
Oligonucleotide synthesis is the chemical synthesis of relatively short fragments of nucleic acids with defined chemical structure . The technique is extremely useful in current laboratory practice because it provides a rapid and inexpensive access to custom-made oligonucleotides of the desired...
by the group of Jesper Wengel in 1998, soon after the first synthesis by the group of Takeshi Imanishi in 1997. The exclusive rights to the LNA technology were secured in 1997 by Exiqon A/S, a Danish biotech company.
LNA nucleotides are used to increase the sensitivity and specificity of expression in DNA microarray
DNA microarray
A DNA microarray is a collection of microscopic DNA spots attached to a solid surface. Scientists use DNA microarrays to measure the expression levels of large numbers of genes simultaneously or to genotype multiple regions of a genome...
s, FISH probes, real-time PCR probes and other molecular biology techniques based on oligonucleotides. For the in situ detection of miRNA
Mirna
Mirna may refer to:geographical entities* Mirna , a river in Istria, Croatia* Mirna , a river in Slovenia, tributary of the river Sava* Mirna , a settlement in the municipality of Mirna in Southeastern Sloveniapeople...
the use of LNA is currently (2005) the only efficient method. A triplet of LNA nucleotides surrounding a single-base mismatch site maximizes LNA probe specificity unless the probe contains the guanine base of G-T mismatch.
Using LNA based oligonucleotides therapeutically is an emerging field in biotechnology
Biotechnology
Biotechnology is a field of applied biology that involves the use of living organisms and bioprocesses in engineering, technology, medicine and other fields requiring bioproducts. Biotechnology also utilizes these products for manufacturing purpose...
. The Danish pharmaceutical company Santaris Pharma a/s
Santaris Pharma a/s
Santaris Pharma A/S is a biopharmaceutical company founded in 2003 in Copenhagen, Denmark with a small branch in San Diego, California that opened in 2009...
owns the sole rights to therapeutic uses of LNA technology, and is now developing a new, LNA based, hepatitis C
Hepatitis C
Hepatitis C is an infectious disease primarily affecting the liver, caused by the hepatitis C virus . The infection is often asymptomatic, but chronic infection can lead to scarring of the liver and ultimately to cirrhosis, which is generally apparent after many years...
drug called miravirsen, targeting miR-122
MiR-122
miR-122 is a miRNA that is conserved between vertebrate species. miR-122 is not present in invertebrates, and no close paralogs of miR-122 have been detected. miR-122 expression is specific to the liver, where it has been implicated as a regulator of fatty-acid metabolism in mouse studies. Reduced...
, which is in Phase II clinical testing as of late 2010.
Benefits of the LNA technology
Some of the benefits of using LNA include:
Applications of the LNA technology
Some proven applications of LNA include:
Other therapeutic and diagnostic applications of LNA technology are in development.