Table 1.
Amplification conditions, oligonucleotide combinations, sequence and amplification fragment of multiplex-PCR for K. variicola identification
| Amplification conditions a | Name of combination primers | Shared unique genes, oligonucleotides and sequence (5′- 3′) of each bacterial specie | |||||
|---|---|---|---|---|---|---|---|
| K. pneumoniae | Amplification fragment (bp) | K. variicola | Amplification fragment (bp) | Klebsiella spp. | Amplification fragment (bp) | ||
| phosphohydrolase | phosphoglycerate mutase | phosphopentane phosphatase (mtnC) | |||||
| 1 | M-PCR-1 | KP888-F: AAGCAAGCCAGAACAGAAAG | 888 | KV770-F: TCCCGAGGTTCACATTTCC | 449 | KmtnC-F: CCGCCGACCTTATCACTAC | 340 |
| KP888-R: ACTTCGGTTTTATCCAGGTC | KV770-R: AGCGGGTGAACGTCGATAC | KmtnC-R: AGCGGGTGAACGTCGATAC | |||||
| transferase (yphG) | N-acetyltransferase | phosphopentane phosphatase (mtnC) | |||||
| 1 | M-PCR-2 | KP878-F: ACCGATAACCAGCCTGACTT | 878 | KV1615-F: ACACAACATTTCAGGCGGCT | 499 | KmtnC-F: CCGCCGACCTTATCACTAC | 340 |
| KP878-R: CTTTCTTCTGCCCACTGTTG | KV1615-R: GGGCGTGGCTTTTTTCATCG | KmtnC-R: AGCGGGTGAACGTCGATAC | |||||
| phosphohydrolase | thiopurine S-methyltransferase | phosphopentane phosphatase (mtnC) | |||||
| 2 | M-PCR-3 | KP888-F: AAGCAAGCCAGAACAGAAAG | 888 | KV1000-F: CTGGGATGTGGCAATGGTG | 438 | KmtnC-F: CCGCCGACCTTATCACTAC | 340 |
| KP888-R: ACTTCGGTTTTATCCAGGTC | KV1000-F: AAACTGCGCCTGCTGTATC | KmtnC-R: AGCGGGTGAACGTCGATAC | |||||
aMultiplex-PCR conditions used under the oligonucleotides combinations. 1: 5pmol/reaction of K. variicola and Klebsiella spp, 25pmol/reaction of K. pneumoniae; 2: 25 pmol/reaction of K. pneumoniae, 5 pmol/reaction of K. variicola and 1 pmol/reaction of Klebsiella spp.