TABLE 3.
Targeted mutagenesis and mutation transfer in B. pseudomallei
| Mutant | Recipient strain | Marker (length of flanking homology [bp])a | Transformation efficiency (no. of colonies/ng/109 cells)
|
No. of confirmed transformants/no. of transformants examinedb | |
|---|---|---|---|---|---|
| Avg | Range | ||||
| Mutants created by PCR fragment mutagenesis | |||||
| Δ(mutS) | 1026b | FRT-tmp-FRT (800) | 6.1 | 6.1 | 2/2 |
| Δ(amrRAB) | 1026b | tmp (800) | 2.4 | 0.2-6.1 | |
| Δ(bpeEF-oprC) | 1026b | tmp (1,500) | 4.9 | 0.6-8.5 | 6/6 |
| Δ(amrRAB) | 1026a | tmp (800) | 1.7 | 0.6-2.3 | |
| Δ(mutS) | 1026a | loxP-tmp-loxP (1,500) | 1.2 | 0.3-2.3 | 4/4 |
| Mutants created by chromosomal DNA transfer | |||||
| Δ(amrRAB) | 1026b | tmp | 23 | 1.2-57 | |
| Δ(bpeEF-oprC) | 1026b | tmp | 6.1 | 3.0-9.2 | 1/2 |
| Δ(mutS) | 1026b | loxP-tmp-loxP | 51 | 51 | 4/4 |
| Δ(amrRAB) | 1026a | tmp | 17 | 4.1-30 | |
| Δ(bpeEF-oprC) | 1026a | tmp | 9.3 | 9.3 | 3/4 |
a
tet, tetracycline resistance; tmp, trimethoprim resistance; loxP, recognition site of Cre recombinase; FRT, recognition site of FLP recombinase.
b
Colonies were verified/examined by PCR tests to determine the generation of the appropriate mutation and the loss of wild-type sequences.