Primers used for MLST of Campylobacter fetus

Genes

The Campylobacter fetus MLST scheme uses internal fragments of the following seven house-keeping genes:

aspA (aspatase)
glnA (glutamine synthetase)
gltA (citrate synthase)
glyA (serine hydroxy methyl transferase)
pgm (phospho glucomutase)
tkt (transketolase)
uncA (ATP synthase alpha subunit)

PCR Amplification and sequencing

The same primers are used for sequences and amplification:

aspA1F 5'-CCT ATG ACT TTA GGT CAA GAG-3'
aspA2F 5'-TGT AGC TAG AGT ACG GCA AG-3'

glnA1F 5'-GAT GGT AGT TCT ATA GAC GC-3'
glnA2F 5'-CTT CCG TTA TCT CCA TAA AGC-3'

gltA1F 5'-CGA TAT AGC GTG GCT AGC TG-3'
gltA2F 5'-AGC GTG AGT AGA TCC TAC G-3'

glyA1F 5'-GAT AAA ATA CTT GGT ATG GAT C-3'
glyA2F 5'-CCC TCT GTT TAT TAA GAC TTC-3'

pgmA1F 5'-AGA GTT GTT TTG GAC GTT GC-3'
pgmA2F 5'-GTA GCT CAT CAA GAG GTC TC-3'

tktA1F 5'-GAG ATA GAT TGG TAT TTA GCG G-3'
tktA2F 5'-GTG ACT ACC TTC TAA ATC TCC-3'

uncA1F 5'-AAG AGT ACG GTG CTA TGG AC-3'
uncA2F 5'-CTC TCA TCA AGA TCG CTT GC-3'

Reaction conditions - PCR

94ºC for 2 min
35 cycles:
94ºC for 20 s
50ºC for 20 s
72ºC for 1 minute
1 cycle:
72ºC for 5 min
4ºC hold

Reaction conditions - sequencing

30 cycles:
96ºC for 10 s
50ºC for 5 s
60ºC for 2 min
4ºC hold