Detection of bacterial (mollicutes) contamination
AssigneeRoche Diagnostics Operations, Inc.
InventorChristian Birkner
Life of this patent
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Abstract
The present disclosure provides a method and system for the PCR amplification of a target sequence which suppresses non-specific amplification products. The disclosure concerns the use of a primer pair optimized to amplify a nucleic acid of a contaminant in the background of genomic DNA of a first organism. When DNA from a second organism suspected for comprising the contaminant is subjected to the same PCR-based amplification reaction, detection sensitivity and specificity of the contaminant is enhanced when an amount of genomic DNA of the first organism is present in the amplification reaction.
Claims21
1-18. (canceled)
A method of detecting a bacterial contaminant within a sample, the method comprising the steps of:
- purifying nucleic acid comprising a sample;
- preparing a reaction mixture including a first primer and a second primer, the first and second primers being specific for a target sequence of nucleic acid comprising a bacterial contaminant within the sample;
- combining the reaction mixture with the nucleic acid purified in said step of purifying;
- combining the reaction mixture with a predetermined amount of a second purified nucleic acid from a second sample, the second purified nucleic acid being free of the bacterial contaminant;
- performing polymerase chain reaction on the reaction mixture combined with the nucleic acid purified in said step of purifying and the second purified nucleic acid, said step of performing amplifying the target sequence comprising the bacterial contaminant;
- and
- detecting presence of the target sequence.
A composition comprising,
- nucleic acid comprising a sample;
- and
- DNA comprising Chinese hamster ovary cells, the DNA comprising the Chinese hamster ovary cells being free of prokaryotic DNA, wherein the sample is selected from the group consisting of amniotic fluid and eukaryotic cells.
A kit comprising:
- a thermostable DNA polymerase;
- purified DNA from a eukaryotic source;
- a first primer;
- and
- a second primer, the first and second primers being specific for amplifying a target sequence of nucleic acid of a prokaryotic source, the purified DNA from the eukaryotic source being free of the target sequence of the prokaryotic source.
Dependencies are inferred from the claim text.