Skip to main content
Journal of Medical Genetics logoLink to Journal of Medical Genetics
. 1995 Feb;32(2):129–130. doi: 10.1136/jmg.32.2.129

Modification of standard proteinase K/phenol method for DNA isolation to improve yield and purity from frozen blood.

N N Ahmad 1, A B Cu-Unjieng 1, L A Donoso 1
PMCID: PMC1050235  PMID: 7760323

Abstract

Research in medical genetics may frequently involve freezing of large numbers of peripheral blood samples. This is a convenient method for storing blood for subsequent DNA isolation and analysis. An area of potential concern is the low yield of DNA from blood samples that have been frozen. Here we report a modification of the widely used standard proteinase K/phenol DNA isolation method for improving the yield and purity of DNA from frozen blood samples, by an initial trypsinisation of whole blood before cell lysis to obtain lymphocytic nuclei and subsequent DNA purification. We report an increased total yield of DNA with pretrypsinised blood as well as improved purity. These results indicate that trypsinisation of thawed whole blood helps the deproteinisation process, reducing the amount of protein associated with the nuclear pellet. This modification to improve yield and purity of DNA from frozen blood samples should be useful to laboratories performing DNA based diagnostic work or studying molecular genetic mechanisms of disease.

Full text

PDF
130

Images in this article

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Beutler E., Gelbart T., Kuhl W. Interference of heparin with the polymerase chain reaction. Biotechniques. 1990 Aug;9(2):166–166. [PubMed] [Google Scholar]
  2. Borst M., Miller D. M. DNA isolation and Southern analysis: a clinician's view. Am J Med Sci. 1990 May;299(5):356–360. doi: 10.1097/00000441-199005000-00011. [DOI] [PubMed] [Google Scholar]
  3. Kunkel L. M., Smith K. D., Boyer S. H., Borgaonkar D. S., Wachtel S. S., Miller O. J., Breg W. R., Jones H. W., Jr, Rary J. M. Analysis of human Y-chromosome-specific reiterated DNA in chromosome variants. Proc Natl Acad Sci U S A. 1977 Mar;74(3):1245–1249. doi: 10.1073/pnas.74.3.1245. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Reinhart W. H., Haeberli A., Stark J., Straub P. W. Influence of blood withdrawal and anticoagulant on clotting activity, hematologic data, and certain rheologic measurements. J Lab Clin Med. 1990 Jan;115(1):98–103. [PubMed] [Google Scholar]
  5. Ross K. S., Haites N. E., Kelly K. F. Repeated freezing and thawing of peripheral blood and DNA in suspension: effects on DNA yield and integrity. J Med Genet. 1990 Sep;27(9):569–570. doi: 10.1136/jmg.27.9.569. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Taş S. Purification of DNA from clotted blood. Clin Chem. 1990 Oct;36(10):1851–1851. [PubMed] [Google Scholar]
  7. Yates J. R., Malcolm S., Read A. P. Guidelines for DNA banking. Report of the Clinical Genetics Society working party on DNA banking. J Med Genet. 1989 Apr;26(4):245–250. doi: 10.1136/jmg.26.4.245. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Medical Genetics are provided here courtesy of BMJ Publishing Group

RESOURCES