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- W2037173619 abstract "A polymerase-chain-reaction-based detection system forMycoplasma fermentanswas established. The highly conservedtufgene, which encodes elongation factor Tu of prokaryotes, served as target sequence for the PCR. With two PCR oligodeoxynucleotides, which were selected fromM. fermentansspecific sequences of thetufgene, we amplified a 850 base pair DNA fragment. Via the biotin-moiety of one primer the PCR fragments were immobilized on streptavidin-coated microtitre plates. After alkaline denaturation a digoxigenin-labelledM. fermentansspecific DNA probe was hybridized to the single stranded immobilized PCR fragment. Detection was performed by addition of an alkaline phosphatase conjugated anti-digoxigenin antibody. 4-methyl-umbelliferyl-phosphate was used as a fluorogenic substrate. Amplification of 10 fg chromosomal target DNA was detected by this «DNA enzyme immuno assay (DEIA)» technique, corresponding to seven genome copies. Our study supports the presumption that thetufgene proves to be a suitable target sequence for the PCR based detection of any bacterial species. Furthermore, hybridization of PCR fragments with radio-labelled DNA probes should no longer be necessary, because a very sensitive non-radioactive test system can easily be established with the «DEIA» technique." @default.
- W2037173619 created "2016-06-24" @default.
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- W2037173619 date "1996-02-01" @default.
- W2037173619 modified "2023-10-10" @default.
- W2037173619 title "Development of an amplification and hybridization assay for the specific and sensitive detection ofMycoplasma fermentansDNA" @default.
- W2037173619 doi "https://doi.org/10.1006/mcpr.1996.0002" @default.
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