Method for detecting live microbiological contaminants in a food product sample
AssigneeCompagnie Gervais Danone
InventorsFrançois Gendre, Pierre Brignon
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Abstract
A process for detecting live microbiological contaminants in a food product sample, according to which the presence of mRNA coding for the synthesis of a protein involved in the synthesis of proteins by the contaminants is detected in the sample.
Claims16
Process for distinguishing live microbiological contaminants from dead microbiological contaminants and detecting live microbiological contaminants in a food product sample, comprising detecting, in the sample, the presence of mRNA coding for a protein involved in the synthesis of proteins by the live contaminants, wherein the presence of mRNA is indicative of a presence of live microbiological contaminants in the sample.
Process for detecting live bacterial contaminants in a food product sample comprising cells that contain mRNA coding for an elongation factor, wherein the process comprises:
- (a) providing a sample of the food product to be tested;
- (b) reverse transcribing mRNA in said sample to produce cDNA;
- (c) amplifying said cDNA by polymerase chain reaction (PCR);
- and
- (d) detecting a coding region of a gene encoding elongation factor EF-Tu using a pair of oligonucleotide primers in the PCR that are specific for said coding region, wherein said pair of oligonucleotide primers is:
- B1:
- 5′ CGTGGAAGGCGACGMRRAG 3′ (SEQ. ID NO. 1)
- B2:
- 5′ CGGAAGTAGAACTGCGGACGGTAG 3′ (SEQ. ID NO. 2).
Process for detecting live yeast contaminants in a food product sample comprising cells that contain mRNA coding for an elongation factor, wherein the process comprises:
- (a) providing a sample of the food product to be tested;
- (b) reverse transcribing mRNA in said sample to produce cDNA;
- (c) amplifying said cDNA by polymerase chain reaction (PCR);
- and
- (d) detecting a coding region of a gene encoding elongation factor EF-1a using a pair of oligonucleotide primers in the PCR that are specific for said coding region, wherein said pair of oligonucleotide primers is:
- L1:
- 5′ TCCATGGTACAAGGGTTGGGAA 3′ (SEQ. ID NO. 3)
- L2:
- 5′ GCGAATCTACCTAATGGTGGGT 3′ (SEQ. ID NO. 4).
Process for detecting live fungal contaminants in a food product sample comprising cells that contain mRNA coding for an elongation factor, wherein the process comprises:
- (a) providing a sample of the food product to be tested;
- (b) reverse transcribing mRNA in said sample to produce cDNA;
- (c) amplifying said cDNA by polymerase chain reaction (PCR);
- and
- (d) detecting a coding region of a gene using a pair of oligonucleotide primers in the PCR that are specific for said coding region, wherein said pair of oligonucleotide primers is:
- M1:
- 5′ GCTGGTATCTCCAAGGATGG 3′ (SEQ. ID NO. 5)
- M2:
- 5′-CGACGGACTTGACTTCRGTGG 3′ (SEQ. ID NO. 6).
Process for detecting live microbiological contaminants in a food product sample comprising cells that contain mRNA coding for an elongation factor, wherein the process comprises:
- (a) providing a sample of the food product to be tested, wherein the food product contains cells;
- (b) lysing said cells;
- (c) reverse transcribing mRNA;
- (d) amplifying the product of step (c) using oligonucleotide primers specific for a coding region of a gene that encodes an elongation factor of a bacteria, yeast, or fungi;
- (e) separating amplified products;
- (f) detecting said amplified products;
- and
- (g) comparing products detected in step (f) with mRNA of a known microorganism;
- wherein said oligonucleotide primers are selected from the group consisting of:
- (1) B1:
- 5′ CGCTGGAAGGCGACGMRRAG 3′ (SEQ. ID NO. 1) and
- B2:
- 5′ CGGAAGTAGAACTGCGGACGGTAG 3′ (SEQ. ID NO. 2)
- (2) L1:
- 5′ TCCATGGTACAAGGGTTGGGAA 3′ (SEQ. ID NO. 3)
- L2:
- 5′ GCGAATCTACCTAATGGTGGGT 3′ (SEQ. ID NO. 4),
- (3) M1:
- 5′ GCTGGTATCTCCAAGGATGG 3′ (SEQ. ID NO. 5), and
- M2:
- 5′-CGACGGACTTGACTTCRGTGG 3′ (SEQ. ID NO. 6).
Process for distinguishing live microbiological contaminants from dead microbiological contaminants and detecting live microbiological contaminants in a food product sample, according to which the presence of mRNA coding for the synthesis of a protein involved in the synthesis of proteins by the live contaminants is detected in the sample by performing RT-PCR technique, wherein said oligonucleotide primers comprise at least two of the primer pairs B1/B2, L1/L2, and M1/M2.
Dependencies are inferred from the claim text.