18. 3 M NaAc prepared with RNase-free water.
19. Autoclaved double-distilled water.
20. RNase-free water.
21. Glycogen.
22. 0.4 M NaCl prepared with RNase-free water.
2.3 Marker Labeling
2.3.1 CIP Treatment of
RNA Oligomers with 5
0
Mono- or Tri-phosphate
1. Markers (Table 1).
2. 1.5 mL Eppendorf tubes and all size tips.
3. Centrifuge.
4. Incubator.
5. CIP (together with Buffer).
6. RNase inhibitor (RNase OUT).
7. RNase-free water.
8. Phenol:chloroform:isoamyl alcohol (25:24:1), pH 4.3.
9. Chloroform:isoamyl alcohol (24:1).
10. Ethanol.
11. 3 M NaAc prepared with RNase-free water.
12. Glycogen.
2.3.2 T4 PNK Treatment
1. Glasses, combs, and clips used for gel preparation.
2. Gel electrophoresis tank and power device.
3. Phosphor screen and Typhoon biomolecular imager.
4. Heat block.
5. Incubator.
6. Blades.
7. 1.5 mL Eppendorf tubes and all size tips (with and without
barriers).
8. Centrifuge.
9. Eppendorf Thermomixer.
10. Needles and syringes.
11. Phosphor screen.
12. Kim wipe.
13. Geiger counter.
14. Markers (Table 1).
15. RNase-free water.
16. T4 PNK (together with PNK buffer).
17. [γ32 P] ATP (3000 Ci/mM).
18. RNase inhibitor (RNase OUT).
Identification and Quantification of sRNAs
231