TABLE 2.
Primers used for gene-walking PCRa
| Primer | Sequence (5′-3′) |
|---|---|
| Arbitrary primers | |
| arb1 | GGCCACGCGTCGACTAGTACNNNNNNNNNNGATAT |
| arb2 | GGCCACGCGTCGACTAGTAC |
| arb6 | GGCCACGCGTCGACTAGTACNNNNNNNNNNACGCC |
| Specific primers | |
| Internal | |
| P101 | CGCACTTTGTACCGCCTCTGTTC |
| P111 | CGACTGCGTTACCAAGTACAGCAC |
| P201 | CAACGAGATGCAACAAACTAAAATC |
| P211 | GGAACAATGAAGAAGGTCATCA |
| External | |
| P102 | GAGGTTGCCGTCGTTTTG |
| P112 | CCTGTTGCTGCCAAAGTGCTG |
| P202 | CCGTAATTTCCAAGCAGAGCATTT |
| P212 | TCATCAGCACTTTGGCAGCAAC |
a
The primer pairs arb1-P101 and arb2-P102 were used in the first gene-walking PCR run, and arb1-P111 and arb2-P112 were used in the second run, to obtain one DNA strand upstream of the cloned sequence b19. arb1-P201 and arb2-P202 were used in the first run, and arb1-P211 and arb2-P212 were used in the second run, to obtain the complementary strand. The primer arb6 was sometimes substituted for arb1 if the latter failed to yield a product.