High-throughput DNA purification with the Qiagen BioRobot™ EZ1J. Chartier, P. Lem, and H.C. Birnboim
DNA Genotek, Ottawa, Ontario, Canada
2006-06-29

The BioRobot™ EZ1 is an automated high-throughput
DNA purification system that uses silica-magneticparticle technology. The EZ1 can purify up tosix Oragene®/saliva samples† in less than 20 minutes,
with excellent DNA recovery.

Different elution volumes
Five Oragene/saliva samples were processed using theEZ1 DNA tissue kit and eluted in volumes of 50, 100,and 200 μL.

DNA analysis

Introduction
Large-scale population studies may involve thecollection of thousands of donor samples. Manualpurification of DNA from these samples can betime- and labor-intensive. A number of manufacturers
offer automated high-throughput DNA purification
systems such as the BioRobot line of machines fromQiagen. The purpose of this study was to determine
the DNA purification performance of the BioRobot
EZ1 System with saliva samples collected with theOragene self-collection kit.

DNA yield was determined by the highly specificFluorescence/DNase method3. The F/D method
quantitates DNA using SYBR Green I™ dye (MolecularProbes) and is more accurate than quantification
by absorbance. DNA purified with each of the threemethods was analyzed by agarose gel electrophoresis
with ethidium bromide staining. The molecularweight of the DNA was determined by comparison
with a Lambda-Hind III digest ladder.

Results

Materials and methods

DNA yield

DNA collection

From 18 Oragene/saliva samples, the median DNA
yield with the EZ1 tissue kit was 3.2 µg per 200 µL ofstarting sample (Figure 1). This was slightly higher
than 2.9 µg obtained with the manual prepIT•L2P
purification protocol. The EZ1 blood kit produced asignificantly lower yield of 1.2 µg.

Oragene kits contain 2 mL of DNA-preserving
solution. The kits were used to collect 2 mL of saliva
from 18 donors (4 mL total volume of Oragene/saliva sample). Prior to purification with theEZ1 System, the Oragene/saliva samples wereincubated overnight at 50°C.

8DNA was extracted from 200 μL of Oragene/saliva
sample using the BioRobot EZ1 System with the EZ1
DNA tissue kit and the EZ1 DNA blood kit (Qiagen).
For both kits, purified DNA was eluted in therecommended volume of 200 μL1. DNA was also
extracted from 200 μL of Oragene/saliva sample
using the manual prepIT™•L2P protocol2.

DNA yield (µg)

DNA purification64
2
0Tissue kit

Blood kit

OrageneFigure 1: Median DNA yield from 200 µL of Oragene/saliva sample
with the EZ1 tissue kit, blood kit, and prepIT purification protocol.

†Saliva samples were collected with Oragene®•DNA or Oragene®•DISCOVER.

(outside the USA)(in the USA)

Some DNA Genotek products may not be available in all geographic regions, contact your sales representative for details.

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Elution volumeMolecular weight of DNA

From five Oragene/saliva samples, the average DNA
yield from 200 µL of starting sample was 2.9 µg withan elution volume of 200 µL. Elution volumes of100 and 50 µL gave lower DNA yields of 2.4 and 2.2 µg,
respectively (Figure 2A). Conversely, the average
DNA concentration increased with decreasing elution
volume: 14.6, 23.7, and 44.6 ng/µL for elution volumesof 200, 100, and 50 µL, respectively (Figure 2B).

All three purification procedures produced DNA
with a molecular weight of 23 kb or higher (Figure 3).

23.1 kb

123M

5ADNA yield (µg)

43Figure 3: Agarose gel electrophoresis of DNA purified with the bloodkit, tissue kit, and prepIT•L2P protocol in Lanes 1, 2 and 3, respectively.
A Lambda-Hind III digest was used as the marker.

2.9
2.4

2

2.2

10200

100

5060
DNA concentration (ng/µL)

B

44.6

40

20Discussion and conclusions
Up to six Oragene/saliva samples may be purified inless than 20 minutes using the BioRobot EZ1 System.
The eluted DNA is of high molecular weight. Formaximum DNA yield, the EZ1 tissue kit is preferredover the blood kit, and the DNA should be eluted
in a volume of 200 µL. Higher throughputs may beobtained using larger BioRobots such as the M48or M96, which use the same MagAttract™ silicamagnetic-particle technology as the EZ1.

23.7
14.6

0200

10050

Elution volume (µL)
Figure 2: Average DNA yields (A) and concentrations (B) with different
elution volumes.

References
123February 2004. EZ1 DNA handbook. Qiagen. Second edition.
Laboratory protocol for manual purification of DNA from 0.5 mL of sample. DNA Genotek. PD-PR-006.
DNA quantification using the Fluorescence/DNase (F/D) assay. Replaced with DNA quantification using SYBR Green I dyeand a micro-plate reader. DNA Genotek. PD-PR-075.

Oragene®•DNA is not available for sale in the United States.
Oragene®•DISCOVER is for research use only, not for use in diagnostic procedures.
®Oragene is a registered trademark and prepIT™ is a trademark of DNA Genotek Inc. All other brands and names contained herein are the property of their respective owners.
All DNA Genotek protocols, white papers and application notes, are available in the support section of our website at www.dnagenotek.com.

MK-AN-006 Issue 3/2011-12© 2011 DNA Genotek Inc., a subsidiary of OraSure Technologies, Inc., all rights reserved.

www.dnagenotek.com • support@dnagenotek.com

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