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US 11,873,534 B2Granted
Method for controlling the quality
AssigneeElectrochaea GmbH
InventorsDoris Hafenbradl, Felix Popp
Life of this patent
Estimating the term…
Abstract
The present invention refers to a method for typing Archaea and quickly discriminating contaminants in pure strain cultures of methanogenic Archaea, leading to isolation of variants with mutations from the culture population.
Claims16
A method for detecting the presence of methanogenic microorganism Methanothermobacter thermoautotrophicus DSM3590, comprising:
- a. obtaining a sample containing one or more methanogenic microorganisms and releasing DNA from the one or more methanogenic microorganisms to receive an unpurified DNA sample or obtaining a sample containing purified DNA of the one or more methanogenic microorganisms;
- b. using the unpurified DNA sample or the purified DNA sample according to step a in a polymerase chain reaction (PCR) amplification using a primer pair comprising a first primer and a second primer, wherein the first primer comprises a nucleotide sequence selected from the group consisting of SEQ ID NO:
- 1, SEQ ID NO:
- 2, and SEQ ID NO:
- 3;
- and the second primer comprises a nucleotide sequence selected from the group consisting of SEQ ID NO:
- 4;
- SEQ ID NO:
- 5, and SEQ ID NO:
- 6, wherein each of the first primers and each of the second primers are capable of hybridizing with the DNA of the methanogenic microorganism Methanothermobacter thermoautotrophicus DSM3590 and wherein the primer pair of SEQ ID NO:
- 1 and SEQ ID NO:
- 4, or SEQ ID NO:
- 2 and SEQ ID NO:
- 5, or SEQ ID NO:
- 3 and SEQ ID NO:
- 6 are used to PCR amplify said DNA, to receive an amplified DNA sequence (amplicon), wherein the amplicon comprises at least one restriction enzyme recognition sequence;
- c. purifying the amplicon;
- d. performing a first digestion reaction with the amplicon of step c by incubating the amplicon in a reaction buffer with at least one first restriction enzyme, which recognizes a first recognition sequence for a time sufficient to form restriction fragments;
- e. determining the number of the formed restriction fragments and their size (restriction pattern) and;
- f. genotyping the one or more methanogenic microorganisms on the basis of the restriction pattern.
17 more elements
A quality control kit for use in detecting the presence of methanogenic microorganism Methanothermobacter thermoautotrophicus DSM3590 within a given sample, the quality control kit comprising:
- at least one container;
- a primer pair used to PCR amplify a DNA sequence of the methanogenic microorganism Methanothermobacter thermoautotrophicus DSM3590 wherein the primer pair comprises:
- a first primer serving as a forward primer and comprising SEQ ID NO:
- 1 and a second primer serving as a reverse primer and comprising SEQ ID NO:
- 4, wherein at least one of the first primer and the second primer is coupled to a detectable marker selected from the group consisting of a radioisotope, a fluorescent compound, an enzyme, and an enzyme co-factor;
- or
- a first primer serving as a forward primer and comprising SEQ ID NO:
- 2 and a second primer serving as a reverse primer and comprising SEQ ID NO:
- 5, wherein at least one of the first primer and the second primer is coupled to a detectable marker selected from the group consisting of a radioisotope, a fluorescent compound, an enzyme, and an enzyme co-factor;
- or
- a first primer serving as a forward primer and comprising SEQ ID NO:
- 3 and a second primer serving as a reverse primer and comprising SEQ ID NO:
- 6, wherein at least one of the first primer and the second primer is coupled to a detectable marker selected from the group consisting of a radioisotope, a fluorescent compound, an enzyme, and an enzyme co-factor;
- at least one first restriction enzyme or at least one first and at least one second restriction enzyme;
- optionally at least one buffer.
12 more elements
Dependencies are inferred from the claim text.