''Nde''I is an

endonuclease Endonucleases are enzymes that cleave the phosphodiester bond within a polynucleotide chain. Some, such as deoxyribonuclease I, cut DNA relatively nonspecifically (without regard to sequence), while many, typically called restriction endonucleases ...

isolated from '' Neisseria denitrificans''. In

molecular biology Molecular biology is the branch of biology that seeks to understand the molecular basis of biological activity in and between cells, including biomolecular synthesis, modification, mechanisms, and interactions. The study of chemical and physi ...

, it is commonly used as a

restriction enzyme A restriction enzyme, restriction endonuclease, REase, ENase or'' restrictase '' is an enzyme that cleaves DNA into fragments at or near specific recognition sites within molecules known as restriction sites. Restriction enzymes are one class o ...

Recognition sequence

Recognition sequence of ''Nde''I: 5'CATATG 3'GTATAC The ends generated by ''Nde''I digest: 5'---CA TATG---3' 3'---GTAT AC---5'

Use in molecular biology

''Nde''I is a specific Type II restriction enzyme that cuts open specific target sequences, unlike

exonucleases Exonucleases are enzymes that work by cleaving nucleotides one at a time from the end (exo) of a polynucleotide chain. A hydrolyzing reaction that breaks phosphodiester bonds at either the 3′ or the 5′ end occurs. Its close relative is th ...

. This enzyme is used in gene cloning to cut open reading frames in the plasmid of certain bacteria such as E. coli and insert a foreign gene, such as the gfpuv gene that codes for bio fluorescence of the jelly fish ''

Aequorea victoria ''Aequorea victoria'', also sometimes called the crystal jelly, is a bioluminescent hydrozoan jellyfish, or hydromedusa, that is found off the west coast of North America. The species is best known as the source of two proteins involved in biolu ...

''. ''Nde''I is useful in generating heterologous DNA construct because it contains the

start codon The start codon is the first codon of a messenger RNA (mRNA) transcript translated by a ribosome. The start codon always codes for methionine in eukaryotes and Archaea and a N-formylmethionine (fMet) in bacteria, mitochondria and plastids. The ...

ATG. It therefore can be used in some

expression vector An expression vector, otherwise known as an expression construct, is usually a plasmid or virus designed for gene expression in cells. The vector is used to introduce a specific gene into a target cell, and can commandeer the cell's mechanism for ...

s, such as those in the pET series of vectors, for the ligation of the start of a gene when making an expression construct. Note however that ''Nde''I generates only a two-base overhang and therefore has a lower melting temperature than other restriction enzymes that generate a four-base overhang. It has a lower

ligation Ligation may refer to: * Ligation (molecular biology), the covalent linking of two ends of DNA or RNA molecules * In medicine, the making of a ligature (tie) * Chemical ligation, the production of peptides from amino acids * Tubal ligation, a meth ...

efficiency, (Note that this paper has an error describing ''Nde''I as a four-base cutter.) since ligation is affected by the ability of the ends to anneal and a two-base overhang has a significantly lower melting temperature compared to a 4-base overhang. Ligation of ''Nde''I-generated ends is therefore best performed with higher ligase concentration with a longer ligation time, whether at room temperature, 14-16^°C, or at 4^°C.

References

Molecular biology Bacterial enzymes Restriction enzymes {{enzyme-stub