TABLE 2.
Oligonucleotide primers used in inverse PCR with template plasmids pYMS4505 and pTM100-ail to mutagenize the yadA and ail genes, respectivelya
| Primer | Sequence (5′-3′)b | 5′-3′ positions | Purpose |
|---|---|---|---|
| yadA-1193r | AAGGCTTTCATGACCAATGG | 1193-1974 | Deletion of aa 193-218 of YadA |
| yadA-1272f | GAAAAAACACAGGAAAATACAAATAAAAG | 1272-1300 | |
| yadA-1310r | CTCAGCTGATCTTTTATTTGTATTTTCCTG | 1310-1281 | Deletion of aa 231-246 of YadA |
| yadA-1356f | GTGCTAGGGATCGCAAATAACTATAC | 1356-1381 | |
| yadA-1355r | GCTAGAAGACTTGTTGTCTGCATACG | 1355-1330 | Deletion of aa 247-262 of YadA |
| yadA-1404f | TTGGAAAATGCGCGTAAAGAGGC | 1404-1426 | |
| yadA-1403r | TGTTTCAGCACTTTTACTATCAGT | 1403-1380 | Deletion of aa 263-277 of YadA |
| yadA-1449f | TTGAATATGGCAAAAGCACACTC | 1449-1471 | |
| yadA-1448r | AACATCTTTAGACTGAGCAAAAGC | 1448-1425 | Deletion of aa 278-292 or 293-307 of YadA |
| yadA-1494f | TTAGAAACTGCTGAAGAACATGC | 1494-1516 | |
| yadA-1493r | AGTTGTTCTAGCAACACTATTTGAG | 1493-1469 | |
| yadA-1538r | AGTTGTTCTGGCAACACTATTTGC | 1538-1515 | Deletion of aa 308-322 of YadA |
| yadA-1584f | TTAGCAAGCGCTAATGTGTATGC | 1584-1606 | |
| yadA-1583r | CGCCTCAGCTGATTTTTTATTTGC | 1583-1560 | Deletion of aa 323-363 of YadA |
| yadA-1707f | ACAGATCATAAATTCCGTCAACTTGAC | 1707-1733 | |
| yadA-1256r | TTGCGCGACATTCACTG | 1256-1240 | Deletion of aa 214-228 of YadA |
| yadA-1302f | TCAGCTGAGCTGTTAGCA | 1302-1319 | |
| yadA-1346r | CTTGTTGTCTGCATACGCAT | 1346-1327 | Deletion of aa 244-258 of YadA |
| yadA-1392f | AGTGCTGAAACATTGGAAAAT | 1392-1412 | |
| yadA-1358r | CACGCTAGAAGACTTGTTGTCTGC | 1358-1335 | Deletion of aa 248-262 of YadA |
| yadA-1404f | TTGGAAAATGCGCGTAAAGAGGC | 1404-1426 | |
| yadA-1370r | TGCGATCCCTAGCACGCT | 1370-1353 | Deletion of aa 252-266 of YadA |
| yadA-1416f | CGTAAAGAGGCTTTTGCTCAGTCTAAA | 1416-1442 | |
| yadA-1382r | AGTATAGTTATTTGCGATCCCTA | 1382-1360 | Deletion of aa 256-270 of YadA |
| yadA-1428f | TTTGCTCAGTCTAAAGATGTT | 1428-1448 | |
| yadA-1391r | TTTACTATCAGTATAGTTATTTGCGAT | 1391-1365 | Deletion of aa 259-273 of YadA |
| yadA-1437f | TCTAAAGATGTTTTGAATATGGCA | 1437-1460 | |
| yadA-1415r | CGCATTTTCCAATGTTTCAGCA | 1415-1394 | Deletion of aa 267-281 of YadA |
| yadA-1461f | AAAGCACACTCAAATAGTGTTGC | 1461-1483 | |
| yadA-1427r | AGCCTCTTTACGCGCAT | 1427-1411 | Deletion of aa 271-285 of YadA |
| yadA-1473f | AATAGTGTTGCTAGAACAACTTTAGAAAC | 1473-1501 | |
| yadA-1481r | AACACTATTTGAGTGTGCTTTTGC | 1481-1458 | Deletion of aa 289-318 of YadA |
| yadA-1572f | TCAGCTGAGGCGTTAGC | 1572-1588 | |
| yadA-1505r | AGCAGTTTCTAAAGTTGTTCTAGCA | 1505-1481 | Deletion of aa 297-326 of YadA |
| yadA-1596f | AATGTGTATGCAGACAGCAAGT | 1596-1617 | |
| yadA-1517r | TGCATGTTCTTCAGCAGTTTCTAA | 1517-1494 | Deletion of aa 301-330 of YadA |
| yadA-1608f | GACAGCAAGTCTTCTCACACAC | 1608-1629 | |
| yadA-1571r | TTTTTTATTTGCATGTTCTTCAGCAGT | 1571-1545 | Deletion of aa 319-333 of YadA |
| yadA-1617f | TCTTCTCACACACTAAAAACTGCA | 1617-1640 | |
| yadA-1616r | CTTGCTGTCTGCATACACATTAG | 1616-1594 | Deletion of aa 334-348 of YadA |
| yadA-1662f | GTAAGTAATTCGACTAAGAAAGCAATCC | 1662-1689 | |
| yadA-1661r | AGTCACATCGGTATAGCTATTTGCA | 1661-1637 | Deletion of aa 349-363 of YadA |
| yadA-1707f | ACAGATCATAAATTCCGTCAACTTGAC | 1707-1733 | |
| yadA-1706r | GTATTGATTCGATTCACGGATTGCTTTC | 1706-1679 | Deletion of aa 364-370 of YadA |
| yadA-1728f | CTTGACAACCGGTTAGATAAACTTGAC | 1728-1754 | |
| yadA-1727r | TTGACGGAATTTATGATCTGTGTATTGA | 1727-1700 | Deletion of aa 371-377 of YadA |
| yadA-1749f | CTTGACACACGAGTTGACAAAG | 1749-1770 | |
| yadA-1748r | TTTATCTAACCGGTTGTCAAGTTGA | 1748-1724 | Deletion of aa 378-384 of YadA |
| yadA-1770f | GGTTTAGCCAGTTCAGCC | 1770-1787 | |
| yadA-1280r | TGTTTTTTCAATTTCTTTCTTTAATTGC | 1280-1253 | Deletion of aa 222-236 of YadA |
| yadA-1326f | AATGCGTATGCAGACAAC | 1326-1343 | |
| yadA-1706r | GTATTGATTCGATTCACGGATTGCTTTC | 1706-1679 | Deletion of aa 364-385 of YadA |
| yadA-1773f | TTAGCCAGTTCAGCCGCTTTAAAC | 1773-1796 | |
| ail-9 | AAGCGCTTGAGGTCAAAGCACAA | 1490-1512 | Cloning of ail gene |
| ail-12 | TTTTCCGGCAGCGGCCCGG | 3201-3183 | |
| H65Ar | ACCAAACTTATTACTGCCATAGAAGAAATCGTAT | 2458-2424 | His65Ala substitution in loop 2 of Ail |
| H65Af | GCAGGTGATCTTGATTACTATTCAGTAACAATGGGGC | 2459-2494 | |
| D67G L68Gr | ACCGTGACCAAACTTATTACTGCCATAGAAG | 2464-2433 | Asp67Gly, Leu68Gly, Asp67Ala, and Leu68Arg substitutions in loop 2 of Ail |
| D67G L68Gf | GGAGGTGATTACTATTCAGTAACAATGGGGCCATCTTTC | 2465-2503 | |
| D67A L68Rf | GCACGAGATTACTATTCAGTAACAATGGGGCCATCTTTC | 2465-2503 | |
| S100r | CTTAACCTTTCCGTGAGCAGCAC | 2563-2540 | Ser100Asp substitution in loop 3 of Ail |
| S100Df | GATTCTGTATTTGATGGGTCAGTCAGTACAAGTAAG | 2564-2599 |