Dideoxynucleotides, or ddNTPs, are nucleotides lacking a 3'-hydroxyl (-OH) group on their deoxyribose sugar. Nucleotides are Organic compounds that consist of three joined structures a nitrogenous base a Sugar, and a Phosphate group Deoxyribose, also known as D-Deoxyribose and 2-deoxyribose, is an Aldopentose &mdash a Monosaccharide containing five Carbon Since deoxyribose already lacks a 2'-OH, dideoxyribose lacks hydroxyl groups at both its 2' and 3' carbons. The lack of this hydroxyl group means that, after being added by a DNA polymerase to a growing nucleotide chain, no further nucleotides can be added as no phosphodiester bond can be created based on the fact that deoxyribonucleoside triphosphates (which are the building blocks of DNA) allow DNA chain synthesis to occur through a condensation reaction between the 5' phosphate (following the cleavage of pyrophospate) of the current nucleotide with the 3' hydroxyl group of the previous nucleotide. A DNA Polymerase is an Enzyme that assists in DNA replication. A phosphodiester bond is a group of strong covalent bonds between the phosphorus atom in a Phosphate group and two other Molecules over two The dideoxyribonucleosides do not possess a 3' hydroxyl group, hence no further chain elongation can occur once this dideoxynucleotide is on the chain. This can lead to the determination of the DNA sequence. Thus, these molecules form the basis of the dideoxy chain-termination method of DNA sequencing, which was developed by Frederick Sanger in 1977. The term DNA sequencing encompasses biochemical methods for determining the order of the Nucleotide bases Adenine, Guanine, Cytosine The term DNA sequencing encompasses biochemical methods for determining the order of the Nucleotide bases Adenine, Guanine, Cytosine Frederick Sanger, OM, CH, CBE, FRS (born 13 August 1918) is an English biochemist and twice [1]
Dideoxynucleotides are useful in the sequencing of DNA in combination with electrophoresis. Electrophoresis is the most well-known electrokinetic phenomenon. A DNA sample that undergoes PCR (polymerase chain reaction) in a mixture containing all four deoxynucleotides and one dideoxynucleotide will produce strands of length equal to the position of each base of the type that complements the type having a dideoxynucleotide present. Nucleotides are Organic compounds that consist of three joined structures a nitrogenous base a Sugar, and a Phosphate group That is, each nucleotide base of that particular type has a probability of being bonded to not a deoxynucleotide but rather a dideoxynucleotide, which ends chain elongation. Thus, if the sample then undergoes electrophoresis, there will be a band present for each length at which the complement of the dideoxynucleotide is present. It is now common to use fluorescent dideoxynucleotides such that each one of the four has a different fluorescence that can be detected by a sequencer; thus only one reaction is needed.
References
- ^ Sanger, F. , Nicklen, S. , and Coulson, A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci, 74(12):5463-7, 1977.
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