Protocol | Published:

Measurement of telomere length by the Southern blot analysis of terminal restriction fragment lengths

Nature Protocols volume 5, pages 15961607 (2010) | Download Citation

Abstract

In this protocol we describe a method to obtain telomere length parameters using Southern blots of terminal restriction fragments (TRFs). We use this approach primarily for epidemiological studies that examine leukocyte telomere length. However, the method can be adapted for telomere length measurements in other cells whose telomere lengths are within its detection boundaries. After extraction, DNA is inspected for integrity, digested, resolved by gel electrophoresis, transferred to a membrane, hybridized with labeled probes and exposed to X-ray film using chemiluminescence. Although precise and highly accurate, the method requires a considerable amount of DNA (3 μg per sample) and it measures both the canonical and noncanonical components of telomeres. The method also provides parameters of telomere length distribution in each DNA sample, which are useful in answering questions beyond those focusing on the mean length of telomeres in a given sample. A skilled technician can measure TRF length in 130 samples per week.

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Acknowledgements

Telomere research at the Center for Human Development and Aging, University of Medicine and Dentistry of New Jersey, New Jersey Medical School was supported by the National Institute of Aging (NIH grants R01AG021593, R01AG20132, R01AG030678 and 5F30AG032858).

Author information

Affiliations

  1. The Center for Human Development and Aging, New Jersey Medical School, University of Medicine and Dentistry of New Jersey, Newark, New Jersey, USA.

    • Masayuki Kimura
    • , Rivka C Stone
    • , Joan Skurnick
    • , Xiaobin Lu
    • , Xiaojian Cao
    •  & Abraham Aviv
  2. Cardiovascular Genetics Division, Department of Internal Medicine, University of Utah School of Medicine, Salt Lake City, Utah, USA.

    • Steven C Hunt
  3. Harley Consulting, Murphys, California, USA.

    • Calvin B Harley

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Contributions

M.K. oversaw the development and refinement of the TRF method, generated data shown in the figures and contributed to the writing of the article. R.S. assembled the elements of the paper, focusing on the presentation and writing of the experimental design and methodology. S.H. participated in the writing of the article, focusing on the statistical component of the introduction. J.S. oversaw the modeling and curve-fitting statistics for the TRF length distribution and contributed to the writing of the article. X.L. and X.C. routinely measured TRF length in our laboratory and provided R.S. with information to construct detailed protocols. C.H., who was a central figure in the development of the quantitative method for measuring the TRF distribution, contributed to the writing of the article. A.A. oversaw all elements of the project and participated in the writing of the article.

Competing interests

C.B.H. holds stock in and consults for Geron Corporation, and is involved in the commercialization of telomere length assays.

Corresponding author

Correspondence to Masayuki Kimura.

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DOI

https://doi.org/10.1038/nprot.2010.124

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