User Manual

Denaturing Nucleic
Acid PAGE
10
10.1 Introduction
Criterion
™
TBE-urea gels are used for separation of small RNA and single-stranded DNA (ssDNA)
fragments. Applications include oligonucleotide analysis, RNase protection assays, and northern
blotting.
10.2 Criterion TBE-Urea Gels
10.2.1 Gel Composition
Gel buffer 89 mM Tris, 89 mM boric acid, 2 mM EDTA, 7 M urea, pH 8.3
Crosslinker 3.3% C
Stacking gel 4% T, 3.3% C
Storage buffer 89 mM Tris, 89 mM boric acid, 2 mM EDTA, pH 8.3
Shelf life ~8 weeks at 2–8°C; expiration date is printed on each cassette
10.2.2 Gel Selection Guide
Gel Percentage Optimum Separation Range
5% 50–1,000 nt
10% 25–300 nt
15% 10–50 nt
10.3 TBE-Urea Buffers
See Appendix B for buffer formulations. Do not adjust pH unless directed to do so.
Running buffer (1x) 89 mM Tris, 89 mM boric acid, 2 mM EDTA
Dilute 100 ml 10x stock (catalog #161-0733) with 900 ml diH
2
O
Sample buffer (5x) 89 mM Tris, 89 mM boric acid, 2 mM EDTA, pH 8.0, 12% Ficoll,
(catalog #161-0768) 0.01% bromophenol blue, 0.02% xylene cyanole FF, 7 M urea
10.4 Sample Preparation
Determine the desired ssDNA or RNA concentration for your sample based on the detection method
used. Dilute 4 parts sample with 1 part sample buffer.
Technical Support: 1-800-4BIORAD • 1-800-424-6723 • www.bio-rad.com 27