Biology Reference
In-Depth Information
3.3 Benzonase
®
Treatment
1. Prepare a working solution (WS) stock of Benzonase
®
in diges-
tion buffer containing 10,000 U/mL just prior to digestion
(
see
Note 14
).
2. Once all the sample is in the feed reservoir (including the vol-
ume inside the filter and connections), measure the volume of
retentate reached.
3. Treat the concentrated virus stock with 100 U/mL of
Benzonase
®
by adding 10 μL of WS/mL of sample into the
feed reservoir. Mix by recirculating sample 3 times for 15 s and
let digestion go for 60 min at room temperature (
see
Note 15
).
A representative example of digestion performance using either
the Midjet
®
or Quixstand
®
systems is shown in Fig.
2
.
4. When the time is over continue with diafiltration process as
described in Subheading
3.2.1
or Subheading
3.2.2
.
Fig. 2
Benzonase
®
digestion. Agarose 1 % gel. Lanes: (M) Molecular weight
marker (GeneRuler
TM
1,000 bp DNA ladder, Fermentas). Retentate (1), permeate
(2), and Benzonase
®
treated retentate (100 U/mL, 1 h at room temperature) (3)
using the Midjet
®
system. Retentate (4), permeate (5), and Benzonase
®
treated
retentate (100 U/mL, 1 h at room temperature) (6) using the Quixstand
®
system
