TABLE 1.
Selected RNA Type–specific protocols for the preparation of RiboMarker sequencing libraries
| RiboMarker Protocols | T[1 + 2] | T[2] | T[1 + 2 + 3 + 4] | T[3 + 4] | T[3] | T[4] |
|---|---|---|---|---|---|---|
| RNA Types in the sample | 1, 2, 3, 4 | 1, 2, 3, 4 | 1, 2, 3, 4 | 1, 2, 3, 4 | 1, 2, 3, 4 | 1, 2, 3, 4 |
| Pretreatment # 1 | — | Rnl1 + Rnl2 ligases (+ATP) | PNK (−ATP) pH 6.0 | PNK, 3′-minus (+ATP) pH 7.6 | Terminator 5′-p -dependent exonuclease | RtcB ligase |
| RNA Types conversion | — | 1 → 0 | 3 → 2 4 → 1 | 2 → 1 3 → 4 |
Degrade 1, 4 |
3 → 0 |
| Pretreatment # 2 | — | — | — | Rnl1 + Rnl2 ligases (+ATP) | PNK, 3′-minus (+ATP) pH 7.6 | PNK, 3′-minus (+ATP) pH 7.6 |
| RNA Types conversion | — | — | — | 1 → 0 | 2 → 1 3 → 4 |
2 → 1 |
| Pretreatment # 3 | — | — | — | PNK (−ATP) pH 6.0 | Rnl1 + Rnl2 ligases (+ATP) | Rnl1 + Rnl2 ligases (+ATP) |
| RNA Types conversion | — | — | — | 4 → 1 | 1 → 0 | 1 → 0 |
| Pretreatment # 4 | — | — | — | — | PNK (−ATP) pH 6.0 | PNK (−ATP) pH 6.0 |
| RNA Types conversion | — | — | — | — | 4 → 1 | 4 → 1 |
| Library preparation protocol | T[1 + 2] | T[1 + 2] | T[1 + 2] | T[1 + 2] | T[1 + 2] | T[1 + 2] |
| RNA Types depleted from library | 3, 4 | 1, 3, 4 | None | 1, 2 | 1, 2, 4 | 1, 2, 3 |
| RNA Types enriched in library | 1, 2 (3′-OH Types) | 2 | 1, 2, 3, 4 (all Types) | 3, 4 (3′-P Types) | 3 | 4 |
