Comparison of DNA purified with Oragene®•DNA and the QIAamp™ Mini Kit
Saliva samples collected with Oragene•DNA may be
purified using the Oragene•DNA purification protocol
or QIAamp DNA Mini Kit (Qiagen). Compared to theOragene•DNA protocol, samples purified with QIAampgave equivalent A260/A280 ratios, but the yield was lower
and the DNA was of lower molecular weight.
Introduction
The Oragene•DNA self-collection kit allows saliva
samples to be collected and stored at room temperature.
The DNA purification process begins as soon as the saliva
mixes with the Oragene•DNA solution. Purification iscompleted in the lab with the Oragene•DNA purification
protocol, which uses an alcohol precipitation procedure
to obtain pure DNA. (ref. 1) QIAamp kits use a silicagel membrane and a bind-wash-elute procedure. Nucleic
acids are adsorbed to the silica membrane in the presenceof high concentrations of chaotropic salts. After a wash
step, DNA is eluted under low-salt conditions (ref. 2).
The purpose of this technical bulletin is to compare
DNA yield, purity, and molecular weight of saliva
samples purified using the Oragene•DNA protocol andthe QIAamp DNA Mini Kit.
Materials and Methods
Saliva collection
Two milliliters of saliva were collected from 8 donors
using the Oragene•DNA kit. In the collection vial, thesaliva was automatically mixed with Oragene•DNApreserving solution. The Oragene•DNA/saliva samples
were stored at room temperature before processing.
Oragene•DNA purification
A 200 μL aliquot from each of the Oragene•DNA/saliva
samples was purified according to the Oragene•DNA
purification protocol. In brief, 8 μL (1/25th vol.) ofOragene•DNA Purifier was added to each aliquot.
The mixture was centrifuged and the supernatant
was transferred to a fresh tube. An equal volume of95% ethanol was added to the clear supernatant. The
precipitated DNA was centrifuged and collected as apellet. The ethanol supernatant was discarded and theDNA pellet was rehydrated in TE buffer.

QIAamp purification
A 200 μL aliquot from each of the Oragene•DNA/saliva samples was purified according to the QIAamppurification protocol (ref. 3). In brief, proteinase K and
200 μL of Buffer AL were added and the sample wasincubated at 56°C. After incubation, 200 μL of 95%ethanol was added. The sample was transferred to aQIAamp spin column and centrifuged. The filtrate wasdiscarded. The column was washed once with Buffer
AW1 and then with Buffer AW2. The DNA was eluted in200 μL of Buffer AE.
Oragene•DNA followed by QIAamp purification
Oragene•DNA purified DNA in TE buffer was added toa QIAamp spin column and centrifuged.
DNA analysis
DNA samples purified with the Oragene•DNA, QIAamp,or Oragene•DNA + QIAamp protocols were quantified
by absorbance. The A260/A280 ratios were determined.
To determine molecular weight, the DNA was run on
a 0.8% agarose gel and compared to a Lamda-Hind III
digest ladder.
Results
The average A260/A280 ratios and DNA yields fromthe Oragene•DNA/saliva samples are summarized inTable 1. The A260/A280 ratios were similar for the three
purification protocols. In contrast, DNA yields werehighest with the Oragene•DNA protocol and lowest
with QIAamp. Some loss of DNA was observed whenthe Oragene•DNA purified DNA was passed through aQIAamp column.
Average DNAyield (µg)

A260/A280

Oragene•DNA

11.2

1.73

QIAamp4.1

1.75

Oragene•DNA+ QIAamp

8.7

1.80

Table 1. Average DNA yield and A260/A280 ratios.

Figure 1 shows the molecular weights of DNA extractedusing the three purification protocols. DNA extractedusing the Oragene•DNA protocol consistently had ahigher molecular weight compared to DNA that had
passed through a QIAamp spin column.
Oragene•DNAQIAamp Oragene•DNA& QIAamp

23.1 kb

Figure 1. Agarose gel electrophoresis of purified DNA.
A Lambda-Hind III digest was used as the marker in lane 1.

Discussion and Conclusions
The A260/A280 ratios of Oragene•DNA and QIAamppurified DNA were similar. This suggests that bothkits are effective at removing contaminants from thesaliva sample. However, DNA yields were lowest with
the QIAamp protocol. This may be because highmolecular-weight DNA elutes slowly from the silicagel membrane. Less DNA loss was observed whenOragene•DNA purified DNA was passed through aQIAamp column. In this case, the DNA was moresolubilized, and less may have been trapped by thecolumn. This trapping effect may also explain thelower-molecular-weight of the QIAamp-purified DNA
as seen by agarose gel electrophoresis. In summary, theOragene•DNA purification protocol gave the highest
yield and the highest molecular weight of DNA.

References
1. Laboratory Protocol for Manual Purification of DNA from 0.5mL ofOragene•DNA/saliva. DNA Genotek. PD-PR-006
2. http://www1.qiagen.com/resources/info/qiagen_purification_technologies_
1.aspx (06/29/04)
3. QIAamp DNA Mini Kit and QIAamp DNA Blood Mini Kit Handbook
Qiagen. September 2001.

© 2004 DNA Genotek
MK-AN-004 Issue 1.3

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