Thermostable uracil DNA glycosylase and methods of use
AssigneeBecton, Dickinson and Company
InventorPaul T. Hamilton
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Abstract
A novel uracil DNA glycosylase enzyme (referred to as Bpa UDG) has been identified in Bacillus pallidus and the gene encoding Bpa UDG has been cloned, sequenced and expressed to produce a recombinant UDG protein. The enzyme is thermostable and exhibits reaction kinetics similar to E. coli UDG. It is effectively inhibited by B. subtilis UGI. Bpa UDG may be used to inactivate contaminating amplicons in nucleic acid amplification reactions, particularly at higher reaction temperatures. It may also be used to generate Bpa UDG-specific antibodies for purification of Bpa-UDG or for detecting Bpa UDG in a sample. Certain Bpa UDG antibodies may inactivate the enzyme and may therefore be useful as substitutes for UGI or heat, or in combination with UGI and/or heat, for controlling UDG activity in a reaction.
Claims20
An isolated nucleic acid sequence selected from the group consisting of SEQ ID NO:5 and nucleic acid sequences encoding SEQ ID NO:6.
Clone ATCC Accession No. 98514.
A method of making a recombinant vector for expression of a uracil DNA glycosylase in a transformed host cell comprising cloning a nucleic acid sequence selected from the group consisting of SEQ ID NO:5 and nucleic acid sequences encoding SEQ ID NO:6 into an expression vector under control of a promoter such that the uracil DNA glycosylase is expressible in the transformed host cell.
Dependencies are inferred from the claim text.