Genevac systems facilitate a wide range of DNA
sample preparation and processing activities, ranging from
straightforward small volume concentration of DNA pellets prior to PCR,
through to high throughput processing of many purified DNA or
oligonucleotide samples. Where a dry sample of DNA is required,
concentration to dryness in a centrifugal evaporator is the ideal
method, presenting the DNA as a dry film in the tube of plate well.
Freeze drying of DNA samples is not recommended because the resultant
DNA powder is easily blown about or moved by electrostatic charges.
DNA Concentration
Concentration of DNA from
samples containing water or a mixture of water and alcohols is a simple
process, which for most researchers requires the miVac DNA concentrator.
Concentration to remove a few hundred microlitres of alcohol from a DNA
pellet takes approximately 10 minutes, or less. This is one of the
simplest applications of a Genevac system – in a miVac DNA concentrator,
set the temperature, select the method for alcohols or water, and press
start.
Oligonucleotides & DNA Purification
The
manufacture of oligonucleotides commonly has two steps where evaporation
is required, evaporation of ammonia solutions following synthesis, and
then subsequent evaporation of purified samples. Protection of
oligonucleotides from damage during drying is critical, especially where
a tag or label has been attached to the DNA. Two studies by Genevac
users show that this can safely be achieved in a Genevac evaporator:
Samples containing ammonia solution should be evaporated using Dri-Pure® (500g)
to prevent sample loss and cross contamination. When ammonia solution
is subjected to a vacuum the ammonia degasses. Degassing must be
controlled or the resultant foam overflows the tube causing sample loss
and cross contamination. In the same way that you control degassing of a
bottle of soda by opening it slowly, gentle application of vacuum is
required, this coupled with the high rotors speeds of Dri-Pure keep every sample where it should be. The latest versions of the EZ-2 come with such a method pre-programmed, and the method is simple to program on the HT-Series.
Where samples have been purified by reverse phase HPLC samples are
presented in a mixture of water and methanol or acetonitrile. The
methods laid out in the Post Purification Sample Handling section should be followed. The imbalance tolerance built into Genevac systems gives researchers confidence that eluates at
different ends of the gradient will still dry down, and if there is too
much imbalance the system will automatically shut down and tell the
user.
Preparation of DNA Microarrays
For certain microarraying techniques concentrated samples of DNA are required. To maintain such a
concentration is difficult due to the volatile nature of the buffers
holding the DNA, therefore samples should be made up to the required
volume immediately prior to analysis. A study by the Wellcome Trust Centre for Human Genetics in Oxford has shown that when an EZ-2 is used in this process the data quality returned by the system is significantly better, reducing the no call rate by 80% or
more.
Useful Papers:
Drying Ammonia Solutions - Application note
Useful Links: