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December 01, 2009 (Vol. 29, No. 21)

Pressure Cycling for Sample Preparation

PCT System Provides Automated Alternative to Manual Methods

  • Genomics

    Click Image To Enlarge +
    Figure 3. Genomic DNA extracted by PCT Shredder is less sheared than DNA extracted by the popular beadmill method.

    DNA was extracted from fresh spinach leaves in DNAzol (Invitrogen, part of Life Technologies) by PCT and by a standard bead-beater protocol. Prior to pressure cycling, spinach was ground for 20 seconds in the PCT shredder. PCT extraction was performed for 30 cycles at 35,000 psi. Bead-beater extraction was performed in a bead beater for 10 10-second pulses with 1 mm zirconia beads.

    Since bead beating generates a large amount of heat, samples were placed on ice between bead-beating pulses to prevent overheating. Following sample disruption by PCT or bead beating, DNA was extracted from the DNAzol as recommended by the manufacturer.

    After DNA purification, triplicate samples were analyzed by electrophoresis. Analysis by gel electrophoresis shows that bead beater-extracted DNA is highly sheared, while PCT-extracted genomic DNA exhibits much less shearing (Figure 3). After extraction by PCT or bead beating, DNA was purified using the DNAzol isolation protocol for plants according to manufacturer’s instructions.

  • Standardization

    The life sciences are increasingly becoming quantitative disciplines that demand a high level of standardization lacking in conventional sample-preparation methods. Traditional sample-preparation methods are generally manual, time consuming, highly variable, and costly in terms of lost opportunity.

    PCT provides rapid, automated, reproducible, and versatile extraction of molecules of interest from almost any sample type, allows the user to exquisitely control the test parameters, and is consequently a method that can effectively bring standardization to sample preparation.

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