17. Xylene cyanol.
18. 0.4 M NaCl prepared with RNase-free water.
19. Ethanol.
2.2 Primer
Purification
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. Syringes and needles.
7. Blades.
8. Tumbling machine.
9. 1.5 mL Eppendorf tubes and all size tips.
10. Centrifuge.
11. Kim wipe.
12. UV lamp and silica gel plates.
13. Nano drop.
14. Primers: these reagents can be customized (Table 1). Noticed
that two popular sRNA library preparation kits (Illumina
®
TruSeq
® Small RNA Library Prep Kit and NEBNext
® Multiplex Small RNA Library Prep Set for Illumina
®
) have different
primer sequences but share the same barcodes and 5
0 adapter
sequence.
15. 1Â TBE buffer.
16. FA buffer: 95% deionized formamide, 0.025% bromophenol
blue, 0.025% xylene cyanol, 5 mM EDTA, 0.025% SDS [20].
17. Urea-polyacrylamide gel reagents.
Table 2
Different concentrated urea-polyacrylamide gel recipe
Components
Concentration
10%
15%
20%
UreaGel buffer
8 mL
8 mL
15 mL
UreaGel concentrate
32 mL
48 mL
120 mL
UreaGel diluent
40 mL
24 mL
15 mL
10% APS
640 μL
640 μL
1.2 mL
TEMED
32 μL
3 2μL
6 0μL
Total volume
80 mL
80 mL
150 mL
230
Di Sun et al.