Modified guanidinium thiocyanate method for human sperm DNA isolation

Amjad Hossain, B. Rizk, A. Behzadian, I. H. Thorneycroft

Research output: Contribution to journalArticle

24 Citations (Scopus)

Abstract

Mammalian sperm chromatin is highly condensed, so isolating DNA from such chromatin can be a formidable task. The procedures that produce high quality DNA from somatic cells fail to yield quality sperm DNA. In this study we have modified the previously used guanidinium method to make it simple and efficient in isolating human sperm DNA. In our method, the lysis buffer contained guanidinium, sodium citrate, sarkosyl, proteinase K and mercaptoethanol. Proteinase K was not used in the original guanidinium method but was included in our protocol. CsCl centrifugation of the lysate, as described in the original procedure, was omitted. Instead, isopropyl alcohol was added directly to the lysis buffer to harvest the DNA. This modified guanidinium method generated high molecular weight DNA while the other two methods resulted in considerable DNA degradation. There was no difficulty in restriction enzyme digestion of DNA prepared by the modified method as revealed by Southern blot analysis. Since the modified guanidinium method is a simple one-step procedure which avoids homogenization, organic solvents, centrifugation and, more importantly, produces degradation-free DNA, it could be the method of choice when DNA from mature germ cells is needed.

Original languageEnglish (US)
Pages (from-to)953-956
Number of pages4
JournalMolecular Human Reproduction
Volume3
Issue number11
StatePublished - 1997
Externally publishedYes

Fingerprint

Spermatozoa
DNA
Guanidine
Endopeptidase K
Centrifugation
Chromatin
Buffers
guanidine thiocyanate
2-Propanol
Mercaptoethanol
DNA Restriction Enzymes
Southern Blotting
Germ Cells
Digestion
Molecular Weight

Keywords

  • Guanidinium method
  • Isolation
  • Sperm DNA

ASJC Scopus subject areas

  • Cell Biology
  • Developmental Biology
  • Genetics
  • Embryology
  • Obstetrics and Gynecology

Cite this

Hossain, A., Rizk, B., Behzadian, A., & Thorneycroft, I. H. (1997). Modified guanidinium thiocyanate method for human sperm DNA isolation. Molecular Human Reproduction, 3(11), 953-956.

Modified guanidinium thiocyanate method for human sperm DNA isolation. / Hossain, Amjad; Rizk, B.; Behzadian, A.; Thorneycroft, I. H.

In: Molecular Human Reproduction, Vol. 3, No. 11, 1997, p. 953-956.

Research output: Contribution to journalArticle

Hossain, A, Rizk, B, Behzadian, A & Thorneycroft, IH 1997, 'Modified guanidinium thiocyanate method for human sperm DNA isolation', Molecular Human Reproduction, vol. 3, no. 11, pp. 953-956.
Hossain, Amjad ; Rizk, B. ; Behzadian, A. ; Thorneycroft, I. H. / Modified guanidinium thiocyanate method for human sperm DNA isolation. In: Molecular Human Reproduction. 1997 ; Vol. 3, No. 11. pp. 953-956.
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