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Primers used for MLST of Streptococcus thermophilus
Genes
The Streptococcus thermophilus MLST scheme uses internal fragments of the following housekeeping genes:
ddlA (D-alanine--D-alanine ligase)
glcK (glucose kinase)
proA (gamma-glutamyl phosphate reductase)
ptsI (phosphoenolpyruvate-protein phosphotransferase)
serB (phosphoserine phosphatase)
tkt (transketolase)
Primers for PCR amplification and sequencing
| Gene | Direction | Primer Sequence |
|---|---|---|
| ddlA | F | TCAAGTGTGGCTATGGA |
| R | GTAGATGGCTCCATCCTC | |
| glcK | F | TGGGCAGAAACTCAAGA |
| R | AACACCACCACCGATAAC | |
| proA | F | CCCGTCTCATCCAAACTGTG |
| R | TGGTTGAAGTTAGGGCCTCA | |
| ptsI | F | ACGTACTAGTCACTCTGCAA |
| R | CACGAAGCAAGGCACGTAAT | |
| serB | F | GGTCCAAGAAGAAGTAATTGA |
| R | GACCTTATACAAATCTGGTT | |
| tkt | F | GCAGCACCAATGGGTTACAC* |
| R | GGAGCACCATGACCAATG |
*(corrected on Nov. 4th, 2019 by removing an erroneous extra 'CA')
PCR conditions
PCR amplification is performed at an annealing temperatures of 54°C for all genes.
Allele templates
ddlA (450 bp):
CTTGAAGTTGCTGGAGTGCCACAGGTTGCCTACACAGGCTACATCGAGGGTGAAGATTTGGAAGCAGCAGTAGCAG
AGACACTTGAAAAATTGACCTTCCCAGTTTTTGTCAAACCAGCCAACATGGGGTCATCTGTTGGGATTTCTAAAGC
TGAAAATGAATCAGAGCTTCGTTCAGCGATTGATCTTGCTCTCAAATATGATAGCCGTATCTTGATTGAGCAAGGT
GTAGTTGCTCGTGAAATCGAAGTTGGTATCCTTGGAAATACGACTGTCAAAACGACTGATCCGGGTGAAGTGGTCA
AAGACGTGGCTTTCTATGATTACCAAGCCAAGTACATTGACAATAAGATTACCATGGATATTCCAGCTCGCGTGCC
TGTCGAAGTCATGACGCAAATGCGTGCCTATGCGGCCAAGGCCTTTCGTGCTCTAGGGGGTTGCGGTCTT
glcK (450 bp):
CTTGGTATTCCATTCGCTATTGATAATGATGCTAATGTGGCTGCACTGGGTGAACGTTGGGTTGGTGCTGGTGCTA
ACAATCGGAATGTTGTCTTTATAACATTGGGTACAGGTGTTGGTGGCGGTGTTATCGCTGATGGTAACTTAATTCA
TGGTGTTGCCGGTGCTGGTGGGGAAATTGGTCACATTATTGTTGAACCTGACACAGGATTTGAGTGTACTTGCGGA
AACAAGGGGTGTCTGGAAACTGTAGCTTCAGCAACAGGTATTGTACGTGTAGCACATCATTTGGCAGAAAAATACG
AAGGAAACTCTTCTATTAAAGCTGCTGTAGACAATGGTGAGTTTGTGACAAGTAAAGATATTATCGTAGCTGCTAC
TGAAGGTGATAAGTTTGCTGACATCATTGTTGATAAAGTCTCTAAATACCTCGGACTTGCAACAGCAAAC
proA (519 bp):
CCAGTCATCGAAACGGGTGTTGGTAACTGTCATATTTATGTGGATAAATATGCTAACTTAGATATGGCGACACAGA
TTGTCATCAATGCCAAGACCCAACGACCAAGTGTGTGTAATGCTGCAGAATCTCTCGTTGTTCATGCTGATATTGT
AGAAGAATTCTTGCCTAACTTGGAAAAAGCTATTTCAAAAATTCAGTCTGTTGAGTTCCGCGCGGATGAAAGGGCT
TTGAAACTTATGGAAAAAGCTGTACCCGCTTTACCAGAGGATTTTGCGACAGAGTTTCTTGACTACATTATGTCTG
TTAAAGTAGTAGACAGCCTTGATGAGGCGATTAATTGGATTAATACTTACACGACATCACATTCAGAAGCTATCGT
GACTCAGGATATCAGTCGTGCTGAGCAATTCCAAGACGATGTTGACGCTGCAGCTGTCTATGTCAATGCCTCGACT
CGTTTCACAGACGGTTTCGTCTTTGGACTGGGTGCTGAAATCGGAATCTCAACCCAAAAAATG
ptsI (501 bp):
GTCCTTGGAATAAACAACATAACTGAACTTGTTAAAGATGGTGACATTTTAGCGGTTTCAGGTATTACCGGTGAAG
TTGTTATTAATCCAACTGAAGAACAAATTGCTGAGTTTAAAGCAGCTGGTGAAGCTTACGCCAAACAAAAGGCAGA
ATGGGCTCAACTTAAAGATGCTCCAACAGTTACTGCTGATGGAAAACACTTCGAGTTAGCAGCAAATATCGGTACA
CCTAAAGACGTTGAAGGTGTTAACGATAATGGTGCAGAAGCAGTTGGTCTTTATCGTACAGAATTCTTGTATATGG
ATTCTCAAGATTTCCCAACCGAAGAAGACCAATATGAAGCATACAAAGCAGTTCTTGAAGGTATGAATGGTAAACC
TGTTGTTGTTCGTACAATGGATATCGGTGGGGATAAAGAACTTCCTTACTTTGACCTTCCTAAAGAAATGAATCCA
TTCTTGGGTTACCGTGCATTACGTATTTCTATCTCTGAAACTGGT
serB (498 bp):
GAGGCTGGTGTTGGTCGAGAGGTCGCTGAAATCACAGAGCGGGCCATGCGTGGAGAATTAGATTTCAGACAGGCTT
TGAATGAGCGTGTGGCAACCCTTAAGGGATTGCCTGACAGTATATTTGAGAAGGTCTATGCCCGCATTCATTTCAA
CAAGGGAGCCAAGGAACTTGTGGATGAACTCCACTCACGTGGCTTCAAGGTAGGTTTAGTTTCTGGTGGTTTCCAT
GAGACAGTTGATAGACTGGCAAAAGAGGCAGGTATTGATTATGTGAAGGCCAATCACCTTGAGGTGATCGATGGTT
TCTTAACTGGAAAGGTTTATGGTGAGATTGTTACCAAAGATGTCAAGGTAGCTAAGCTTAAAGATTGGGCAGCGGA
AAATGGACTTAAGCTCTCTCAAACCATTGCTATGGGTGATGGAGCCAATGATTTGCCTATGATTAAGACAGCGGGT
ATAGGCATCGCCTTTTGTGCCAAACCAATTGTCCGTGTACAA
tkt (525 bp):
CGTTTTGTTTTGTCAGCAGGTCATGGTTCAATGCTTCTTTATGCGCTCCTTCATTTGTCAGGATTTAAAGATTTGA
GCATTGAAGAGCTTAAACAATTCCGTCAATGGGGTTCTAAGACACCTGGTCACCCAGAATTTGGCCATACAGTTGG
TGTAGATGCTACGTCAGGTCCACTTGGGCAAGGTATTGCTATGGCTGTCGGTATGGCACAAGCTGAACGTTTCTTG
GCTTCTCGATACAACAAGGAAGGCTTCCCAATCTTTGACCATTATACTTATGTTATTGCGGGAGATGGTTGCTTTA
TGGAAGGCGTTTCGGCCGAAGCATCATCTTACGCTGGTCTTCAAAAATTGGATAAATTGATTGTCTTGTACGATTC
AAATGATATCAATCTTGACGGTGAAACTAAAGATTCATTTACTGAAGATGTACGTGCACGTTATGAAGCTTATGGT
TGGAACACTGAATTTGTTCAAGATGGAACAGATATTGAAGCTATCAACGCTGCTATTGAAAGTGCAAAA