Linked Life Data

18438658

Source:http://linkedlifedata.com/resource/pubmed/id/18438658

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
4
pubmed:dateCreated
2008-5-30
pubmed:databankReference
pubmed:abstractText
In this study, we found that deoxyinosine triphosphate (dITP) could inhibit polymerase chain reaction (PCR) amplification of various family B-type DNA polymerases, and 0.93% dITP was spontaneously generated from deoxyadenosine triphosphate during PCR amplification. Thus, it was hypothesized that the generated dITP might have negative effect on PCR amplification of family B-type DNA polymerases. To overcome the inhibitory effect of dITP during PCR amplification, a dITP pyrophosphatase (dITPase) from Thermococcus onnurineus NA1 was applied to PCR amplification. Genomic analysis of the hyperthermophilic archaeon T. onnurineus NA1 revealed the presence of a 555-bp open reading frame with 48% similarity to HAM1-like dITPase from Methanocaldococcus jannaschii DSM2661 (NP_247195). The dITPase-encoding gene was cloned and expressed in Escherichia coli. The purified protein hydrolyzed dITP, not deoxyuridine triphosphate. Addition of the purified protein to PCR reactions using DNA polymerases from T. onnurineus NA1 and Pyrococcus furiosus significantly increased product yield, overcoming the inhibitory effect of dITP. This study shows the first representation that removing dITP using a dITPase enhances the PCR amplification yield of family B-type DNA polymerase.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jun
pubmed:issn
0175-7598
pubmed:author
pubmed:issnType
Print
pubmed:volume
79
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
571-8
pubmed:meshHeading
pubmed:year
2008
pubmed:articleTitle
Characterization of a dITPase from the hyperthermophilic archaeon Thermococcus onnurineus NA1 and its application in PCR amplification.
pubmed:affiliation
Korea Ocean Research and Development Institute, Ansan, South Korea.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't