TABLE 1.
Strains and oligonucleotides
| Sulfolobus strain or oligonucleotide | Genotype | Source or derivation | Sequence (5′ to 3′) |
|---|---|---|---|
| Strains | |||
| MR31 | pyrE131 (bp 159–177 deleted) | Spontaneous Foar mutant (49) | |
| DG185 | Wild type | ATCC 33909 | |
| SD67 | Δogg::pyrESso pyrE131 | This study (see Materials and Methods) | |
| SD67-7 | Δogg::pyrE mutant pyrE131 | Spontaneous Foar mutant of SD67 | |
| RJ4 | Δdbh::pyrEStoV4 | This study (see Materials and Methods) | |
| RJ12 | wild-type | MR31 transformed to Pyr+ by pSAPE5a | |
| RJ13 | Δogg::pyrE mutant pyrE131 | Spontaneous Foar mutant of SD67 | |
| RJ8 | Δogg Δdbh | RJ13 transformed by PCR product of RJ4 | |
| RJ401 | Δdbh::pyrEStoV4 | Spontaneous Foar mutant of RJ4 | |
| RJ803 | Δogg Δdbh | RJ80 transformed to Pyr+ by pSAPE5a | |
| Oligonucleotidesb | |||
| 5′ anchor | ACTTACAAGCAATAAATGAGGCAAATGGAACGCCCCCAGTAACAACTCCCAATATCATAT | ||
| 3′ anchor | P-ATGTCGACTGCAGAACTAACGACGAATGAAAATATGTCAGGATGGTTGGGGAGTTTCCTT | ||
| Control insert | P-CAAATTGAGCNCCTTTGACT | ||
| oxoG insert | P-CAAATTGAGC[8oG]CCTTTGACT | ||
| Downstream scaffold | GCTCAATTTGATATGATATTG | ||
| Upstream scaffold | AGTCGACATAGTCAAAGG | ||
| delSso Sa1367f | AGTTAAGAAAAAAACCAGTAAGAAAAAGAAGAGAAAGAAAAGTGCTAAGATGTCTTAATCTCACAAAG | ||
| delSso Sa1367f | TCACAACTAATCATTTATACTCCTTTTTATACTTTTTTGTTCTATTTTACGATATGAGAGAGGTTTATC | ||
| delSto Sa0554f | CCTTAAATGCTTATACACCAAATACTAAATGTAAATGATAGTGGCAGTGGGTATTTAAAC | ||
| delSto Sa0554r | TTAAGCAAAATCCTTAACTCGTTGCAATTAAATGTCGAAGAAATCCCACTGCCTAGGTCT |
a
E. coli cloning plasmid containing the wild-type pyrE gene (53).
b
Synthetic DNAs used in strain constructions and TLS assays (see Materials and Methods).