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US 10,538,818 B2Granted
Methods and kits to determine the sensitivity of strains of Lactococcus lactis bacteria to phage infection
AssigneeDsm IP Assets B.V.
InventorsDouwe Van Sinderen, Finn Vogensen, Jennifer Mahony, Witold Kot, Stuart Ainsworth
Life of this patent
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Abstract
A kit useful for determining the phage susceptibility of one or more strains of Lactococcus lactis is described. The disclosure also describes methods for formulation of mixed defined starter cultures using strains from different phage sensitivity groups.
Claims15
A method comprising:
- (i) providing a sample comprising at least one strain of Lactococcus lactis (L. lactis);
- (ii) performing a PCR analysis of the sample using at least a first primer pair adapted to generate a test amplicon correlating to a region of the cell wall polysaccharide (cwps) operon that is not conserved among strains of L. lactis, and using a control primer pair adapted to generate a control amplicon correlating to a region of the cwps operon that is conserved amongst strains of L. lactis;
- and
- (iii) detecting the presence or absence of the test amplicon in the strain of L. lactis, and
- (iv) detecting the presence of the control amplicon in the strain of L. lactis,
- wherein the test amplicon comprises a sequence selected from the group consisting of SEQ ID NOs:
- 9 to 11 and 13 to 15, and wherein the control amplicon comprises SEQ ID NO:
- 12.
5 more elements
A method comprising:
- (i) providing a sample comprising at least one strain of Lactococcus lactis (L. lactis);
- (ii) performing a multiplex PCR analysis of the sample using
- (a) a first primer pair adapted to generate a first amplicon, wherein the first amplicon comprises SEQ ID NO:
- 9 or SEQ ID NO:
- 13;
- (b) a second primer pair adapted to generate a second amplicon, wherein the second amplicon comprises SEQ ID NO:
- 10 or SEQ ID NO:
- 14;
- (c) a third primer pair adapted to generate a third amplicon, wherein the third amplicon comprises SEQ ID NO:
- 11 or SEQ ID NO:
- 15;
- and
- (d) optionally, a control primer pair adapted to amplify a sequence conserved in L. lactis;
- and
- (iii) detecting the presence or absence of the amplicons.
12 more elements
A method comprising:
- (i) providing a sample comprising at least one strain of Lactococcus lactis (L. lactis);
- (ii) performing a multiplex PCR analysis of the sample using
- (a) a first primer pair adapted to generate a first amplicon correlating to a region of the cwps operon, wherein the first primer pair comprises a forward primer comprising SEQ ID NO:
- 1 and a reverse primer comprising SEQ ID NO:
- 2;
- (b) a second primer pair adapted to generate a second amplicon correlating to a region of the cwps operon, wherein the second primer pair comprises a forward primer comprising SEQ ID NO:
- 3 and a reverse primer comprising SEQ ID NO:
- 4;
- (c) a third primer pair adapted to generate a third amplicon correlating to a region of the cwps operon, wherein the third primer pair comprises a forward primer comprising SEQ ID NO:
- 5 and a reverse primer comprising SEQ ID NO:
- 6;
- and
- (d) optionally, a control primer pair adapted to amplify a sequence conserved in L. lactis;
- and
- (iii) detecting the presence or absence of the first, second and third amplicons wherein the presence of the first amplicon indicates the presence of a strain that is phage-sensitive, the presence of the third amplicon indicates the presence of a strain that is phage-insensitive, and the presence of the second amplicon indicates the presence of a strain having intermediate phage sensitivity.
12 more elements
A method comprising:
- (i) providing a sample comprising a mix of Lactococcus lactis (L. lactis) strains;
- (ii) performing a multiplex PCR analysis of the sample using
- (a) a first primer pair adapted to generate a first amplicon correlating to a region of the cwps operon that is present in L. lactis strains MG1363 and SK11 but is not present in L. lactis strains IL1403, KF147, UC509.9 and CV56;
- (b) a second primer pair adapted to generate a second amplicon correlating to a region of the cwps operon that is present in L. lactis strains IL1403 and KF147 but is not present in L. lactis strains MG1363, SK11, UC509.9 and CV56;
- (c) a third primer pair adapted to generate a third amplicon correlating to a region of the cwps operon that is present in L. lactis strains UC509.9 and CV56 but is not present in L. lactis strains MG1363, SK11, IL403, and KF147;
- and
- (d) optionally, a control primer pair adapted to amplify a sequence conserved in L. lactis,
- wherein the PCR analysis provides a plurality of amplicons of different sizes;
- and
- (iii) detecting the level of the first, second and/or third amplicons.
7 more elements
Dependencies are inferred from the claim text.