TABLE 1.
Bacteria and plasmids used in this work
| Bacterial strain or plasmid | Description | Source or reference |
|---|---|---|
| Strains | ||
| Y. enterocolitica | ||
| 6471/76-c (YeO3-c) | Virulence plasmid cured derivative of YeO3 | 47 |
| YeO3-c-Ail | Δail::Km-GenBlock; derivative of YeO3-c; Kmr | 4 |
| YeO3-c-Ail-OCR | Spontaneous OC mutant derivative of YeO3-c-Ail-R; Kmr | 4 |
| E. coli | ||
| JM109 | reaA1 Δlac-pro endA1 gyrA96 thi-1 hsdR17 supE44 relA1 F′ traD36 proAB+lacIqZΔM15 | 56 |
| DH10B | F−mcrA Δ(mrr-hsd RMS-mcrBC), φ80lacZΔM15 ΔlacX74 deoR recA1 endA1 araΔ139Δ(ara leu)7697 galU galK λ−rpsL nupG λ−tonA | Life Technologies |
| HB101/pRK2013 | Triparental conjugation helper strain; Kmr | 12 |
| Plasmids | ||
| pYMS4505 | yadA gene of YeO3 cloned as a 1,735-bp PCR fragment (nucleotides 478-2213)a into the BamHI site of pTM100; Clmr; the yadA gene is transcribed by the constitutive tet promoter of pTM100 | 10 |
| pYMS4505:ΔNECK | Deletion of bp 1194-1271 of the yadA gene in pYMS 4505; expresses YadA with a 26-aa deletion (aa 193-218) | This work |
| pYMS4505:A | Deletion of bp 1257-1301 of the yadA gene in pYMS 4505; expresses YadA with a 15-aa deletion (aa 214-228) | This work |
| pYMS4505:S | Deletion of bp 1281-1325 of the yadA gene in pYMS 4505; expresses YadA with a 15-aa deletion (aa 222-236) | This work |
| pYMS4505:Δ#1 | Deletion of bp 1311-1355 of the yadA gene in pYMS 4505; expresses YadA with a 16-aa deletion (aa 231-246) | This work |
| pYMS4505:B | Deletion of bp 1347-1391 of the yadA gene in pYMS 4505; expresses YadA with a 15-aa deletion (aa 244-258) | This work |
| pYMS4505:Δ#2 | Deletion of bp 1356-1403 of the yadA gene in pYMS 4505; expresses YadA with a 16-aa deletion (aa 247-262) | This work |
| pYMS4505:C | Deletion of bp 1359-1403 of the yadA gene in pYMS 4505; does not express YadA due to a frameshift mutation introduced during construction | This work |
| pYMS4505:D | Deletion of bp 1371-1415 of the yadA gene in pYMS 4505; expresses YadA with a 15-aa deletion (aa 252-266) | This work |
| pYMS4505:E | Deletion of bp 1383-1427 of the yadA gene in pYMS 4505; expresses YadA with a 15-aa deletion (aa 256-270) | This work |
| pYMS4505:Δ#3+6 | Deletion of bp 1398-1448 of the yadA gene in pYMS 4505; expresses YadA with a 17-aa deletion (aa 261-277) | This work |
| pYMS4505:F | Deletion of bp 1392-1436 of the yadA gene in pYMS 4505; expresses YadA with a 15-aa deletion (aa 259-273) | This work |
| pYMS4505:Δ#3 | Deletion of bp 1404-1448 of the yadA gene in pYMS 4505; expresses YadA with a 15-aa deletion (aa 263-277) | This work |
| pYMS4505:H | Deletion of bp 1416-1460 of the yadA gene in pYMS 4505; expresses YadA with a 15-aa deletion (aa 267-281) | This work |
| pYMS4505:I | Deletion of bp 1428-1472 of the yadA gene in pYMS 4505; expresses YadA with a 15-aa deletion (aa 271-285) | This work |
| pYMS4505:Δ#4 | Deletion of bp 1449-1493 of the yadA gene in pYMS 4505; expresses YadA with a 15-aa deletion (aa 278-292) | This work |
| pYMS4505:J | Deletion of bp 1482-1571 of the yadA gene in pYMS 4505; expresses YadA with a 30-aa deletion (aa 289-318) | This work |
| pYMS4505:Δ#5 | Deletion of bp 1494-1538 of the yadA gene in pYMS 4505; expresses YadA with a 15-aa deletion (aa 293-307) | This work |
| pYMS4505:Δ#5-6 | Deletion of bp 1494-1583 of the yadA gene in pYMS 4505; expresses YadA with a 30-aa deletion (aa 293-322) | This work |
| pYMS4505:K | Deletion of bp 1506-1595 of the yadA gene in pYMS 4505; expresses YadA with a 30-aa deletion (aa 297-326) | This work |
| pYMS4505:L | Deletion of bp 1518-1607 of the yadA gene in pYMS 4505; expresses YadA with a 30-aa deletion (aa 301-330) | This work |
| pYMS4505:Δ#6 | Deletion of bp 1539-1583 of the yadA gene in pYMS 4505; expresses YadA with a 15-aa deletion (aa 308-322) | This work |
| pYMS4505:Δ#6+3 | Deletion of bp 1539-1586 of the yadA gene in pYMS 4505; expresses YadA with a 16-aa deletion (aa 308-323) | This work |
| pYMS4505:M | Deletion of bp 1572-1616 of the yadA gene in pYMS 4505; expresses YadA with a 15-aa deletion (aa 319-333) | This work |
| pYMS4505:Δ#7-9 | Deletion of bp 1584-1706 of the yadA gene in pYMS 4505; expresses YadA with a 41-aa deletion (aa 323-363) | This work |
| pYMS4505:N | Deletion of bp 1617-1661 of the yadA gene in pYMS 4505; expresses YadA with a 15-aa deletion (aa 334-348) | This work |
| pYMS4505:O | Deletion of bp 1662-1706 of the yadA gene in pYMS 4505; expresses YadA with a 15-aa deletion (aa 349-363) | This work |
| pYMS4505:P | Deletion of bp 1707-1727 of the yadA gene in pYMS 4505; expresses YadA with a 7-aa deletion (aa 364-370) | This work |
| pYMS4505:Δ1-3 | Deletion of bp 1707-1772 of the yadA gene in pYMS 4505; does not express surface-located YadA; 22-aa deletion (aa 364-385) | This work |
| pYMS4505:Q | Deletion of bp 1728-1748 of the yadA gene in pYMS 4505; expresses YadA with a 7-aa deletion (aa 371-377) | This work |
| pYMS4505:R | Deletion of bp 1749-1769 of the yadA gene in pYMS 4505; expresses YadA with a 7-aa deletion (aa 378-384) | This work |
| pTM100 | Mobilizable derivative of pACYC184 | 31 |
| pTM100-ail | ail gene cloned in a 1,711-bp PCR fragment (nucleotides 1490-3201)b into EcoRV site of pTM100; Clmr | This work |
| pTM100-ail1 | CAC at bp 2459-2461 substituted for GCA in pTM100-ail; expresses Ail with His65Ala substitution (loop 2) | This work |
| pTM100-ail2 | GATCTT at bp 2465-2470 substituted for GGAGGT in pTM100-ail; expresses very low levels of Ail with Asp67Gly and Leu68Gly substitutions (loop 2) | This work |
| pTM100-ail3 | GATCTT at bp 2465-2470 substituted for GCACGA in pTM100-ail; expresses very low levels of Ail with Asp67Ala and Leu68Arg substitutions (loop 2) | This work |
| pTM100-ail4 | TCA at bp 2564-2566 substituted for GAT in pTM100-ail; expresses Ail with Ser100 substituted for Asp (loop 3) | This work |
a
In all yadA mutants, nucleotide positions refer to the numbering of the 2,551-bp sequence containing the yadA gene of Y. enterocolitica O:3 under GenBank accession no. X13882.
b
In all ail mutants, nucleotide positions refer to the numbering of the 3,865-bp sequence containing the ail gene of Y. enterocolitica O:3 under GenBank accession no. AJ605740.