PF8
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Primers used for MLST of Pantoea agglomerans
 

- Genes

The Pantoea agglomerans MLST scheme uses internal fragments of the following six housekeeping genes:

  • fusA (protein chain elongation factor EF-G)
  • gyrB (DNA gyrase)
  • leuS (leucyl-tRNA synthetase)
  • pyrG (CTP synthase)
  • rplB (50S ribosomal protein L2)
  • rpoB (RNA polymerase subunit B)

- Primers for PCR amplification

FusA : F : FusA3 : CATCGGCATCGGTATCAGTGCKCACATCGA
FusA : R : FusA4 : CAGCATCGCCTGAACRCCTTTGTT

gyrB : F : gyrB3 : GCGTAAGCGCCCGGGTATGTA
gyrB : R : gyrB4 : CCGTCGACGTCCGCATCGGTCAT

leuS : F : leuS3 : CAGACCGTGCTGGCCAACGARCARGT
leuS : R : leuS4 : CGGCGCGCCCCARTARCGCT

pyrG : F : pyrG3 : GGGGTCGTATCCTCTCTGGGTAAAGG
pyrG : R : pyrG4 : GGAACGGCAGGGATTCGATATCNCCKA

rplB : F : rplB3 : CAGTTGTTGAACGTCTTGAGTACGATCC
rplB : R : rplB4 : CACCACCACCATGYGGGTGRTC

rpoB : F : Vic3 : GGCGAAATGGCWGAGAACCA
rpoB : R : Vic2 : GAGTCTTCGAAGTTGTAACC

- PCR conditions

PCR amplification is performed at two annealing temperatures of 60°C during 10 cycles and 50°C during 21 cycles for fusA, gyrB, leuS, pyrG and rplB genes, and at an annealing temperatures of 50°C for rpoB.

- Sequencing primers

We use the PCR primers also for sequencing, except for gene gyrB for which primers gyrB3i and gyrB4i are used .

gyrB : F : gyrB3i : AACGCWATCGACGAAGC
gyrB : R : gyrB4i : TGGAAVCCRTCRTTCCAC

- Allele templates

fusA(633 bp)

GGTGTAAACCACAAAATCGGTGAAGTACATGACGGCGCAGCAACCATGGACTGGATGGAGCAGGAACAGGAACGTG GTATTACCATCACCTCCGCTGCGACCACCTGTTACTGGTCTGGTATGGCCAAGCAGTTTGAACCACACCACGTAAA CATTATCGACACCCCAGGACACGTTGACTTCACCATCGAAGTTGAACGTTCTATGCGTGTGCTTGATGGCGCGGTA ATGGTTTACTGTGCAGTTGGTGGCGTTCAGCCACAGTCTGAGACCGTATGGCGTCAGGCTAACAAATATAAAGTTC CACGCATTGCGTTCGTTAACAAAATGGACCGCATGGGTGCTAACTTCCTGAAAGTTGTTAAACAGATGGAAGTTCG TCTGGGCGCAACTCCAGTTCCTCTGCAGCTGGCTATCGGCGCAGAAGAGAAATTCACCGGTGTTGTTGACCTGGTG AAAATGAAAGCAATCAACTGGAACGACGCAGACCAGGGTGTAACCTTCGTCTACGAAGATATCCCAGCTGATATGC AGGAACTGGCTGAAGAATGGCATGCGAAGCTGGTTGAAGCCGCTGCTGAAGGCTCTGATGAGCTGATGGAGAAATT CTTCGGTGGTGAAGAACTGACTGAA

gyrB (417 bp)

GAGGGCATTTCTGCCGCTGAAGTGATCATGACCGTGCTGCATGCTGGCGGTAAGTTCGATGATAACTCCTATAAAG TCTCCGGCGGCCTGCACGGCGTGGGCGTCTCCGTGGTTAACGCCCTGTCGCAGAAGCTGGAGCTGACCATTCGTCG CGAAGGCAAAGTGCATCAGCAGATTTACGTCCATGGCGTACCGCAGGCACCACTGGCGGTGACGGGTGAGACCGAC ATTACCGGGACCCGTGTGCGTTTCTGGCCAAGCCACGAAACCTTTACCAACGTGCGCGATTTTGAGTATGACATCC TGGCGAAACGCCTGCGTGAACTCTCATTCCTGAACTCCGGCGTGTCGATTCGTCTGGATGATAAGCGTGATGGTAA AAGTGATCACTTCCACTATGAAGGCGGCATCCGGGCG

leuS (642 bp)

CAGTGGTTTATCAAAATCACCGATTATGCGGAAGAGCTGCTGAACGACCTGGATACGCTGGAAGAGTGGCCTGAGCA GGTCAAAACCATGCAGCGTAACTGGATTGGCCGCTCTGAAGGTGTCGAAATCACCTTCGATGTGGCTGACAGCGAAG AGAAGGTCACTGTCTACACCACCCGCCCGGATACCTTTATCGGCGCAACCTATGTTGCCGTTGCCGCAGGCCATCCG CTGGCGACCCAGGCGTCGGTCAATAATCCGGCGCTGGCTGACTTTATCGCTGAATGTCGCAATACCAAAGTGGCTGA AGCCGATATGGCGACCATGGAGAAGAAAGGCATGGCGACCGGCCTGTCTGTTGTTCATCCGCTGACCGGCGAGCTTA TCCCGGTCTGGGTTGCCAACTTCGTACTGATGGAATATGGCACCGGCGCTGTGATGGCGGTTCCGGCGCACGATCAG CGCGACTGGGAGTTTGCCACTAAATATGATCTGCCGATTAAGCCGGTCATCCTGAATCTGGATGGTTCCATTCCTGA TGTCAGCATTGCGGCGATGACTGACAAAGGTGCGCTGTTCAACTCAGGTGAGTTTAACGGCATGGATCACGCCACCG GTTTTAACGCCATCGCGGATTCGCTG

pyrG (306 bp)

ATTCTCGAAGCCCGTGGTCTGAACGTGACCATCATGAAGCTGGACCCGTATATCAACGTGGATCCAGGCACCATGAG CCCGACGCAGCACGGTGAAGTTTTCGTCACTGATGACGGCGCTGAAACCGATCTGGATTTGGGTCACTACGAGCGCT TCATCCGTACCAAAATGTCGCGTCGTAACAACTTCACCACTGGCCGTATCTACTCAGAAGTCCTGCGCAAAGAGCGC CGTGGCGACTATCTGGGCGCAACCATTCAGGTTATTCCGCACATCACCAACGCCATCAAAGAACGCATTATTGAA

rplB (333 bp)

CGTTACATCCTGGCCCCTAAAGGCCTGAAAGCCGGCGACCAGATCCAATCTGGCGTTGATGCTGCGATTAAAGCAGG TAACACTCTGCCGATGCGTAACATCCCGGTGGGTTCAACCGTGCATAACGTAGAAATGAAACCAGGCAAAGGCGGTC AGATTGCTCGCTCAGCCGGTGCTTACGTGCAGATCGTTGCGCGTGAAGGTTCTTACGTTACCCTGCGTCTGCGTTCA GGTGAAATGCGTAAAGTCGAGTCTGACTGCCGCGCAACACTGGGCGAAGTCGGCAACGCTGAGCATATGCTGCGCGT TCTGGGTAAAGCTGGTGCAGCCCGT

rpoB (501 bp)

ACGCACTACGGTCGTGTCTGCCCAATCGAAACCCCTGAAGGCCCGAACATCGGTCTGATCAACTCCTTGTCTGTGTAT GCACAGACCAACGAGTACGGTTTCCTTGAGACGCCTTACCGCCGTGTTATCGACGGTCTGGTTTCTGACGAGATTCAC TACCTCTCTGCTATCGAAGAGGGTAACTACGTTATCGCTCAGGCGAACGCCCCTCTTGATGATAACGGCGCGTTTGTT GATGACCTGGTCACCTGCCGTAGCAAAGGCGAATCAAGTCTGTTCAGCCGCGATCAGGTTGACTACATGGACGTTTCG ACTCAACAGATCGTTTCTGTCGGTGCGTCACTGATCCCGTTCCTGGAACACGATGATGCTAACCGCGCCCTGATGGGT GCGAACATGCAACGTCAGGCCGTTCCGACTCTGCGTGCTGATAAGCCGCTGGTTGGTACGGGTATGGAACGTGCTGTT GCCGTTGACTCCGGTGTAACTGCCGTTGCCAAA
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Databases are maintained by Sylvain Brisse using software developed by Keith Jolley at Oxford (Jolley et al. 2004, BMC Bioinformatics, 5:86). Web managers : Ghislaine GUIGON and Jean-michel THIBERGE

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