AL
Basic workflow Figure 2 shows a simplified view of the key processes often carried out
in molecular biology laboratories. Biological samples can include blood, plant material, bacteria or the product of cloning experiments. In each case, DNA is isolated from these samples. When the amount of DNA is too small for additional work, it is amplified by PCR. This DNA can be sequenced or ligated into DNA vectors as part of the cloning process. These vectors are incorporated into modified bacteria (transformation) or eukaryotic cells (transfection), and typically contain reporter genes such
Table 1 – DNA/RNA/protein isolation Procedure
Lysis: Chemically induced destruction of the cell, as well as micelles and pro- teins that protect the nucleic acids.
Adsorption: Specialized magnetic beads or membranes selectively bind nucleic acids. The rest of the mixture obtained in the lysis step (lysate) is discarded.
Washing: A series of wash buffers are then added to remove everything but the desired nucleic acids.
Elution: Elution buffers are added to remove the pure nucleic acids from the magnetic beads or the membrane.
as those coding for fluorescence or antibiotic resistance and which are used to identify successful clones. The desired clones are then selected and grown in bioreactors to produce biological samples that often need to go through the cycle again.
Why automation? Automation of laboratory work is usually justified because it reduces
human error, facilitates better allocation of human resources, provides increased precision and reproducibility, enables automatic digital storage of results and offers greater throughput.
Automation approach(es)
Addition of lysis buffers (basically fatty acid chelators), followed by incubation with mild shaking.
The above mixture is combined with magnetic beads or passed through filter plates. Magnets placed under the sample allow the lysate to be aspirated and discarded. When filter plates are used, pressure or vacuum removes the lysate from the sample.
Depending on the required throughput, a pipettor or dispenser can add the buffers and the previous adsorption step is repeated.
When filtration is used, a collection plate is inserted below the filter plate to collect the purified product.
Tool(s)
• Automated pipettor • Liquid dispenser* • Heating/cooling nest • Shaking nest**
• Automated pipettor • Liquid dispenser* • Pressure-based nest** • Magnetic bead nest** • Vacuum nest**
• Automated pipettor • Liquid dispenser* • Plate washer* • Shaking nest** • Heating/cooling nest**
• Automated pipettor • Pressure-based nest • Magnetic bead nest** • Vacuum-based nest** • Liquid dispenser*
*This equipment is generally optional, but significantly increases the throughput of the process. **The need for this equipment depends on the specific protocol.
Table 2 – PCR: amplify small amounts of DNA to make it easier (possible) to study Procedure
Automation approach(es)
PCR preparation: Creation of a mixture of the template DNA and the necessary reagents.
Thermocycling: An iterative series of denatur- ing and annealing steps.
PCR purification: Isolation of pure DNA from the PCR reaction mixture.
A series of liquid-handling steps add the various reagents to the DNA sample.
The sample plate is transferred to the thermocycler and a predefined program is run.
A series of liquid-handling steps add the various reagents to obtain the purified DNA sample.
Tool(s)
• Automated pipettor • Liquid dispenser
• Thermocycler
• Automated pipettor • Liquid dispenser • Magnetic bead nest • Pressure-based nest
AMERICAN LABORATORY 41 MAY 2016
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