Sister Chromatid Exchange

Jane Bayani1, Jeremy A. Squire1

1 Princess Margaret Hospital and The Ontario Cancer Institute, Toronto, Ontario
Publication Name:  Current Protocols in Cell Biology
Unit Number:  Unit 22.7
DOI:  10.1002/0471143030.cb2207s25
Online Posting Date:  January, 2005
GO TO THE FULL TEXT: PDF or HTML at Wiley Online Library

Abstract

Sister chromatid exchange (SCE) refers to the interchange of DNA between replication products. The technique for detecting such exchanges takes advantage of the semiconservative nature of DNA synthesis. 5′‐bromodeoxyuridine (BrdU) is incorporated into the newly synthesized DNA. Using standard culturing techniques, Colcemid is added to the culture and conventional cytogenetic preparations are made. Differential staining with Hoechst dye and Giemsa allows the newly synthesized DNA within a chromatid to be recognized, since BrdU incorporation results in much weaker staining. SCEs represent a point of DNA template exchange during strand synthesis, visualized as asymmetric chromatid staining or “harlequin” chromosomes.

Keywords: sister‐chromatid exchange; SCE; BrdU; DNA replication

     
 
GO TO THE FULL PROTOCOL:
PDF or HTML at Wiley Online Library

Table of Contents

  • Strategic Planning
  • Basic Protocol 1: Hoechst‐Giemsa Staining for SCE and Replication Banding
  • Reagents and Solutions
  • Commentary
  • Literature Cited
  • Figures
     
 
GO TO THE FULL PROTOCOL:
PDF or HTML at Wiley Online Library

Materials

Basic Protocol 1: Hoechst‐Giemsa Staining for SCE and Replication Banding

  Materials
  • Subconfluent cell culture
  • 5′‐bromodeoxyuridine (BrdU; Sigma): follow manufacturer's instructions and keep away from direct light
  • 10 µg/ml Colcemid (Invitrogen) in H 2O
  • Phosphate‐buffered saline (PBS; appendix 2A)
  • Hoechst 33258 working solution (see recipe)
  • 4% (v/v) Giemsa staining solution
  • 2× SSC (see unit 18.6 for 20×)
  • 15‐ml conical centrifuge tubes
  • Coplin jars
  • Large petri dish
  • UV light source, e.g., the UV light in a tissue culture hood
  • Additional reagents and equipment for harvesting cells and preparing metaphase spreads (unit 22.2)
GO TO THE FULL PROTOCOL:
PDF or HTML at Wiley Online Library

Figures

  •   FigureFigure 22.7.1 Sister chromatid exchange in a human CML spread and Chinese hamster ovary cells. Examples of sister chromatid exchange in a human CML spread (A) and in Chinese hamster ovary cells (B). Images kindly provided by Turid Knutsen, Hesed Padilla‐Nash, and Thomas Ried, Genetics Branch, National Institute of Health, Bethesda, Maryland.

Videos

Literature Cited

Literature Cited
   Barch, M.J. 1991. The ACT Cytogenetics Laboratory Manual, 2nd Edition. Raven Press, New York.
   Ellis, N.A., Proytcheva, M., Sanz, M.M., Ye, T.Z., and German, J. 1999. Transfection of BLM into cultured Bloom syndrome cells reduces the sister‐chromatid exchange rate toward normal. Am. J. Hum. Genet. 65:1368‐1374.
   Gebhart, E. 1981. Sister chromatid exchange (SCE) and structural chromosome aberration in mutagenicity testing. Hum. Genet. 58:235‐254.
   Taylor, J.H., Woods, P.S., and Hughes, W.L. 1957. The organization and duplication of chromosomes as revealed by autoradiographic studies using tritium‐labeled thymidine. Proc. Natl. Acad. Sci. U.S.A. 43:122‐128.
GO TO THE FULL PROTOCOL:
PDF or HTML at Wiley Online Library