How are Oligonucleotides Used
Oligo Primers and Probes
The typical use for oligonucleotides involves synthesizing a nucleotide sequence that is paired or 'reverse-complimentary' to a larger, target nucleic acid strand. There are two general types of use for oligos. As primers, oligos initiate a chemical chain reaction on a target sequence. As an example, the polymerase chain reaction (PCR) uses 2’-deoxyribonucleotides as primers and in this application, small sequences of DNA are augmented.
Oligo primers are frequently used to enable sequence elongation, thus forming a complementary strand to a target DNA sequence. This method is known as dideoxy sequencing, or the Sanger method, and is used for PCR and early forms of DNA sequencing. The function of the oligo primer is to provide the sequence-extending DNA polymerase enzyme with a template on which to operate. This forms a complementary DNA sequence and the end of the process is a double strand DNA molecule.
Primers are created by linking specific nucleotides. This process is carefully controlled to ensure that the resultant molecule is well matched to the target strand. In order to have chain lengthening, it is critical that the primer’s 3’ deoxyribose carbon end coincides with sequences on the template molecule. Applications for oligo primers include DNA sequencing, gene expression, cloning and molecular diagnostics.
Oligos are used as probes as well. In this application, they bind to and hold a target sequence. Applications using oligo probes include blotting procedures such as northern blotting (for RNA) or southern blotting (for DNA), as fluorophore-conjugated sequences in microarrays that detect changes in genes expression or used in screening for genetic diseases or to identify specific pathogens.
In therapeutic applications, antisense oligonucleotides (ASO) take advantage of natural biology and facilitate gene inhibition or gene silencing (destruction) of undesirable or over-active RNA sequences, this in turn blocks expression of certain damaged or overactive proteins which may be causing or facilitating disease.