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Characterization of Effect of Repeated Freeze and Thaw Cycles on Stability of Genomic DNA Using Pulsed Field Gel Electrophoresis

  1. Author:
    Shao, W.
    Khin, S.
    Kopp, W. C.

  2. Author Address

    [Shao, Wen; Khin, Sonny; Kopp, William C.] NCI, Appl Dev Res Directorate, SAIC Frederick, Frederick, MD 21701 USA.;Shao, W (reprint author), NCI Frederick, Appl Dev Res Directorate, SAIC Frederick, POB 358, Clarksburg, MD 20871 USA;wenmd@yahoo.com
    1. Year: 2012
    2. Date: Feb
  2. Journal: Biopreservation and Biobanking
    1. 10
    2. 1
    3. Pages: 4-11
  4. Type of Article: Article
  5. ISSN: 1947-5535
  1. Abstract:

    In this study, we systematically investigated the effects of repeated cycles of freeze/thaw on stability of genomic Deoxyribonucleic acid (DNA) samples as evaluated by changes in DNA size, concentration, and freeze/thaw protocols. The DNA was isolated using standard extraction procedures including phenol/chloroform and commercially available Gentra Puregene and Qiagen QIAmp kits. Changes in DNA were monitored over 18 cycles of freezing and thawing utilizing several freeze/thaw protocols. DNA samples from multiple subjects prepared from whole blood samples were examined by pulsed field gel electrophoresis (PFGE), and shown to have different average molecular sizes and size distribution patterns depending on the extraction method. Results of freeze/thaw experiments, analyzed by PFGE, showed progressive DNA degradation of the samples, with DNA sizes larger than 100 kb most sensitive to freeze/thaw degradation. Increasing the DNA concentration of stored samples from 10 mu g/mL to 100 mu g/mL had a somewhat protective effect on DNA stability. Variations in freeze/thaw protocols did not have a significant impact on DNA stability during repeated freeze/thaw cycles. At freeze/thaw cycle 18, average molecular size and size distribution of all DNA samples tested approached 25 kb, regardless of their initial average size and size distributions. This study provides insight on DNA degradation during freeze/thaw cycles and offers guidance to storage and handling of DNA samples.

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External Sources

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  2. DOI: 10.1089/bio.2011.0016
  3. View record in Web of Science®
  4. WOS: 000300540800003

Library Notes

  1. Fiscal Year: FY2011-2012
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