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.
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. Full details are in this application note on drying ammonia solutions.
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 HPLC Fractions 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, such as the Sequenom MassArray
system 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 MassArray system is significantly better, reducing the no call rate by 80% or more.
Sequenom & MassArray are registred trademarks of Sequenom Inc.