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Humans host complex microbial communities believed to contribute to health maintenance and, when in imbalance, to the development of diseases. Determining the microbial composition in patients and healthy controls may thus provide novel therapeutic targets. For this purpose, high-throughput, cost-effective methods for microbiota characterization are needed.(More)
A cDNA encoding the precursor of the bovine mitochondrial phosphate carrier protein has been cloned from a bovine cDNA library using a mixture of 128 different 17-mer oligonucleotides as hybridisation probe. The protein has an N-terminal extension of 49 amino acids not present in the mature protein. This extension has a net positive charge and is presumed(More)
The Pyrosequencing technology was used for identification of different clinically relevant fungi. The tests were performed on amplicons derived from the 18S rRNA gene using polymerase chain reaction (PCR) universal primers for amplification. Sequencing was performed up to 40 bases in a variable region with a designed general sequencing primer and the(More)
An approach for real-time DNA sequencing without the need for electrophoresis has been developed. The approach relies on the detection of DNA polymerase activity by an enzymatic luminometric inorganic pyrophosphate (PPi) detection assay (ELIDA) (Nyrén, P. (1987) Anal. Biochem. 167, 235-238). The PPi formed in the DNA polymerase reaction is converted to ATP(More)
There is a growing demand for high-throughput methods for analysis of single-nucleotide polymorphic (SNP) positions. Here, we have evaluated a novel sequencing approach, pyrosequencing, for such purposes. Pyrosequencing is a sequencing-by-synthesis method in which a cascade of enzymatic reactions yields detectable light, which is proportional to(More)
The amount of F1-ATPase in chromatophores from Rhodospirillum rubrum was determined by Western blotting using anti-RrF1 rabbit antibodies. 9.1 mmol F1 (mol bacteriochlorophyll)-1 was obtained or 14% of the total protein content of the chromatophores. The turnover rate of the F0F1-ATPase was 17 molecules ATP s-1 during synthesis, 2 molecules ATP s-1 during(More)
A sensitive method for the analysis of inorganic pyrophosphate (PPi) which utilizes the enzymes ATP sulfurylase and firefly luciferase is described. The assay is based on continuous monitoring of the ATP formed in the ATP sulfurylase reaction using purified firefly luciferase. The assay can be completed in less than 2 s and is not affected by inorganic(More)
The possibility of using a new bioluminometric DNA sequencing technique, called pyrosequencing, for typing of human papillomaviruses (HPV) was investigated. A blinded pyrosequencing test was performed on an HPV test panel of 67 GP5+/GP6+ PCR-derived amplification products. The 67 clinical DNA samples were sequenced up to 25 bases and sequences were searched(More)
ATP-diphosphohydrolase (apyrase) catalyzes the hydrolysis of phosphoanhydride bonds of nucleoside tri- and di-phosphates in the presence of divalent cations. This enzyme has broad substrate specificity for nucleotides, which makes it an ideal enzyme for different biotechnical applications, such as DNA sequencing and platelet-aggregation inhibition. The only(More)