TABLE 4.
Numbers of 16S rRNA gene-derived tags and the phylogenetic level at which they are able to discriminate the 140 sequenced bacterial genomes
| Identifier and tag location | Enzyme | Recognition sequence | No. of tags at the level ofa:
|
No. of nonidentified organisms | ||||||
|---|---|---|---|---|---|---|---|---|---|---|
| Domain | Phylum | Class | Order | Family | Genus | Species | ||||
| 16S rRNA gene, | HpyCH4 IV | ACGT | 0 | 1 (NE)c | 0 | 0 | 2 (6) | 19 (10) | 328 (120) | 4 |
| upstream | Sau3AI | GATC | 2 (8) | 3 (12) | 5 (8) | 1 (NE) | 1 (NE) | 17 (8) | 233 (104) | 0 |
| BamHI | GGATCC | 0 | 0 | 0 | 0 | 1 (2) | 5 (5) | 101 (62) | 71 | |
| Csp6I | GTAC | 0 | 0 | 0 | 0 | 2 (NE) | 33 (9) | 374 (129) | 2 | |
| 16S rRNA gene, | HpyCH4 IV | ACGT | 4 (45) | 0 | 1 (2) | 0 | 2 (5) | 10 (23) | 73 (65) | 0 |
| downstream | Sau3AI | GATC | 5 (7) | 6 (7) | 3 (3) | 0 | 3 (6) | 14 (24) | 126 (93) | 0 |
| BamHI | GGATCC | 0 | 0 | 0 | 0 | 0 | 6 (14) | 26 (22) | 104 | |
| Csp6I | GTAC | 1 (2) | 0 | 3 (10) | 5 (19) | 3 (6) | 13 (25) | 83 (78) | 0 | |
| SARST, internalb | V1 region | 0 | 0 | 2 (3) | 1 (4) | 1 (2) | 9 (16) | 162 (124) | 0 | |
Tags were generated in silico with an MmeI-containing linker cassette from the first position of the anchoring enzyme located upstream or downstream of the 27R primer annealing site. Data are presented for HpyCH4IV, Sau3AI, BamHI, and Csp6I as anchoring enzymes. Numbers in parentheses indicate the number of species that can be identified by SP-GSTs at a given phylogenetic level. Since the 16S rRNA gene often has multiple copies per species, tag numbers at a phylogenetic level can be higher than the number of species distinguished at that level. Tags located at more than 3,000 nucleotides from the primer annealing sites were discarded, as a result of which some organisms were not identified.
SARST data for the V1 hypervariable region, which was also present in nine Archaea, are presented as comparison.
NE, no effect. At this level the tag had no effect on the final identification of the species, as additional tags were generated from the same species that allowed for identification at a lower phylogenetic level.