.
1 July 2019
-2-
E-Base
™
DNA electrophoresis protocol
Important guidelines
·
Use 10−200 ng DNA per band for samples with one unique band or up to 500 ng per lane for samples with multiple bands.
·
Dilute samples with high salt concentrations (>50 mM NaCl, >100 mM KCl, >10 mM acetate ions, >10 mM EDTA) 2- to 5-fold in deionized water, TE, or 1X E-Gel
™
Sample Loading
Buffer, in a final volume of 15 μL (48-well gels) or 20 uL (96-well gels).
·
Load E-Gel
™
agarose gels within 30 minutes after opening the pouch; run gels within 1−3 minutes after loading samples.
Step
Action
1–5 min
1
Prepare samples
Prepare DNA samples in deionized water OR 1X E-Gel
™
Sample Loading Buffer.
·
For optimal separation use 20−100 ng of DNA per band for samples with one
unique band or
up to 500 ng per lane for samples with multiple bands.
·
The total sample volume for 48-well gels is 15 μL.
·
The total sample volume for 96-well gels is 20 μL.
5–10 min
2
Prepare gel
cassette
a. Plug the Mother E-Base
™
Device into an electrical outlet.
b. Remove the gel from the package and gently remove the comb(s) from the E-Gel
™
cassette.
c. Insert the cassette into the E-Base
™
Device, starting from the right edge. When properly
inserted, the device indicates its initialized status with a steady red light.
Note
:The protocol type on the display shows EG for E-Gel
™
DNA cassettes, and EP for
E PAGE
™
cassettes.
3
Load samples
Load samples with a multichannel pipettor.
Load a volume of
15 μL in each well for 48-well gels
.
Load a volume of
20 μL in each well for 96-well gels
.
a. Load prepared samples into sample wells. Keep all sample volumes uniform.
b. Load prepared E-Gel
™
DNA ladder into marker wells.
c. Load 1X E-Gel
™
Sample Loading Buffer or deionized water in all empty wells. The buffer for
empty wells should have a similar salt concentration to adjecent sample wells.
