Mechanical disruption of biological samples for nucleic acid extraction

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The following abstract was presented at the Plant, Animal & Microbe Genomes X Conference, January, San Diego, CA, January 2002. If you have any questions, please feel free to contact us at info@btc-bti.com.

 

Evaluation of methods for the high throughput mechanical disruption of biological samples for nucleic acid extraction

Arthur Piorkowski, Gita Vaghani, William Fithian, and David Burden

The extraction of nucleic acids from biological samples may require mechanical disruption methods instead of chemical or enzymatic lysis. This is true for many microbes and solid matrix samples such as seeds or bones. Traditional vortexing or "bead beating" cells using a mixer/amalgamator where the sample is agitated vigorously with glass beads is laborious and inconsistent. To overcome these limitations, we have investigated the use of mechanical grinding mills designed to process samples in 96-well plate format. Initially two different mills were examined for sample disruption, namely the SPEX CertiPrep Model Geno/Grinder 2000 and the Retsch Model MM300 Mixer Mill. These were tested in multiple formats on Bacillus, Saccharomyces, Pichia, and Aspergillus. Both mills were effective in disrupting fungal cells cultured in deep well plates. The effectiveness of lysis was related to the diameter of beads used and motion of the mill. The vertical action of the Geno/Grinder consistently resulted in greater cellular lysis. There was no significant lysis of bacterial cells by either mill. When cells were lysed in standard 96-well plates using the Geno/Grinder, lysis approached 100%. Throughput for microbial cultures was 768 samples in 5 minutes for the Geno/Grinder and 192 for the Retsch Mixer Mill. The Geno/Grinder was also evaluated for the disruption of corn kernels and soybeans. Seeds were effectively homogenized by first soaking in TE buffer followed by grinding with steel balls. Extracted DNA was successfully amplified by PCR. This tool is also being used to disrupt animal tissue, including bone fragments.

The Geno/Grinder provides better sample grinding than other mixer mills for homogenizing soybean, kernels, microbes and more.
Homogenized corn kernels

 

Both the HT Homogenizer and Geno Grinder are effective at cracking yeast in microwell plates

Disrupted yeast cells