Abstract
Single-stranded DNA (ssDNA) intermediates play an important role in processes such as DNA replication and homologous recombination, DNA damage responses, and DNA repair. Using quantitative amplification of ssDNA (QAOS), ssDNA arising during various cellular processes in complex genomes can be quantified at numerous single-copy and repetitive loci. QAOS is a useful tool to gain insights into the cellular processes that involve ssDNA and the roles of proteins in regulating ssDNA production and responses to ssDNA.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Maringele L, Lydall D (2002) EXO1-dependent single-stranded DNA at telomeres activates subsets of DNA damage and spindle checkpoint pathways in budding yeast yku70Delta mutants. Genes Dev 16:1919–1933
Niu H, Chung WH, Zhu Z, Kwon Y, Zhao W, Chi P, Prakash R, Seong C, Liu D, Lu L, Ira G, Sung P (2010) Mechanism of the ATP-dependent DNA end-resection machinery from Saccharomyces cerevisiae. Nature 467:108–111
Zou L, Elledge SJ (2003) Sensing DNA damage through ATRIP recognition of RPA-ssDNA complexes. Science 300:1542–1548
Bertuch AA, Lundblad V (2003) The Ku heterodimer performs separable activities at double-strand breaks and chromosome termini. Mol Cell Biol 23:8202–8215
Grandin N, Damon C, Charbonneau M (2001) Ten1 functions in telomere end protection and length regulation in association with Stn1 and Cdc13. EMBO J 20:1173–1183
Dewar JM, Lydall D (2010) Pif1- and Exo1-dependent nucleases coordinate checkpoint activation following telomere uncapping. EMBO J 29:4020–4034
Foster SS, Zubko MK, Guillard S, Lydall D (2006) MRX protects telomeric DNA at uncapped telomeres of budding yeast cdc13-1 mutants. DNA Repair (Amst) 5:840–851
Booth C, Griffith E, Brady G, Lydall D (2001) Quantitative amplification of single-stranded DNA (QAOS) demonstrates that cdc13-1 mutants generate ssDNA in a telomere to centromere direction. Nucleic Acids Res 29:4414–4422
Pitt CW, Cooper JP (2010) Pot1 inactivation leads to rampant telomere resection and loss in one cell cycle. Nucleic Acids Res 38:6968–6975
Addinall SG, Holstein E-M, Lawless C, Yu M, Chapman K, Banks P, Ngo H-P, Maringele L, Taschuk M, Young A, Ciesiolka A, Lister AL, Wipat A, Wilkinson DJ, Lydall D (2011) Quantitative fitness analysis shows that NMD proteins and many other protein complexes suppress or enhance distinct telomere cap defects. PLoS Genet 4:e1001362
Lee K, Zhang Y, Lee SE (2008) Saccharomyces cerevisiae ATM orthologue suppresses break-induced chromosome translocations. Nature 454:543–546
Negrini S, Ribaud V, Bianchi A, Shore D (2007) DNA breaks are masked by multiple Rap1 binding in yeast: implications for telomere capping and telomerase regulation. Genes Dev 21:292–302
Ngo HP, Lydall D (2010) Survival and growth of yeast without telomere capping by Cdc13 in the absence of Sgs1, Exo1, and Rad9. PLoS Genet 6:e1001072
Jia X, Weinert T, Lydall D (2004) Mec1 and Rad53 inhibit formation of single-stranded DNA at telomeres of Saccharomyces cerevisiae cdc13-1 mutants. Genetics 166:753–764
Lazzaro F, Sapountzi V, Granata M, Pellicioli A, Vaze M, Haber JE, Plevani P, Lydall D, Muzi-Falconi M (2008) Histone methyltransferase Dot1 and Rad9 inhibit single-stranded DNA accumulation at DSBs and uncapped telomeres. EMBO J 27:1502–1512
Zubko MK, Guillard S, Lydall D (2004) Exo1 and Rad24 differentially regulate generation of ssDNA at telomeres of Saccharomyces cerevisiae cdc13-1 mutants. Genetics 168:103–115
Zubko MK, Maringele L, Foster SS, Lydall D (2006) Detecting repair intermediates in vivo: effects of DNA damage response genes on single-stranded DNA accumulation at uncapped telomeres in budding yeast. Methods Enzymol 409:285–300
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2012 Springer Science+Business Media New York
About this protocol
Cite this protocol
Holstein, EM., Lydall, D. (2012). Quantitative Amplification of Single-Stranded DNA. In: Bjergbæk, L. (eds) DNA Repair Protocols. Methods in Molecular Biology, vol 920. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-998-3_23
Download citation
DOI: https://doi.org/10.1007/978-1-61779-998-3_23
Published:
Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-1-61779-997-6
Online ISBN: 978-1-61779-998-3
eBook Packages: Springer Protocols