Abstract
This protocol describes the use of a 16plex PCR for the purpose assessing DNA quality after isothermal whole genome amplification (WGA). In short, DNA products, generated by amplification multiple displacement amplification, are forwarded to PCR targeting 15 short tandem repeats (STR) as well as amelogenin generating up to 32 different PCR products. After amplification, the PCR products are separated via capillary electrophoresis and analyzed based on the obtained DNA profiles. Isothermal WGA products of good DNA quality will result in DNA profiles with efficiencies of >90 % of the full DNA profile.
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References
Van Loo P, Voet T (2014) Single cell analysis of cancer genomes. Curr Opin Genet Dev 24:82–91. doi:10.1016/j.gde.2013.12.004
Kroneis T, Geigl JB, El-Heliebi A, Auer M, Ulz P, Schwarzbraun T, Dohr G, Sedlmayr P (2011) Combined molecular genetic and cytogenetic analysis from single cells after isothermal whole-genome amplification. Clin Chem 57(7):1032–1041
van Beers EH, Joosse SA, Ligtenberg MJ, Fles R, Hogervorst FB, Verhoef S, Nederlof PM (2006) A multiplex PCR predictor for aCGH success of FFPE samples. Br J Cancer 94(2):333–337. doi:10.1038/sj.bjc.6602889
El-Heliebi A, Kroneis T, Zohrer E, Haybaeck J, Fischereder K, Kampel-Kettner K, Zigeuner R, Pock H, Riedl R, Stauber R, Geigl JB, Huppertz B, Sedlmayr P, Lackner C (2013) Are morphological criteria sufficient for the identification of circulating tumor cells in renal cancer? J Transl Med 11:214. doi:10.1186/1479-5876-11-214
El-Heliebi A, Kroneis T, Wagner K, Meditz K, Kolb D, Feichtinger J, Thallinger GG, Quehenberger F, Liegl-Atzwanger B, Rinner B (2014) Resolving tumor heterogeneity: genes involved in chordoma cell development identified by low-template analysis of morphologically distinct cells. PLoS One 9(2):e87663. doi:10.1371/journal.pone.0087663
Panaro NJ, Yuen PK, Sakazume T, Fortina P, Kricka LJ, Wilding P (2000) Evaluation of DNA fragment sizing and quantification by the agilent 2100 bioanalyzer. Clin Chem 46(11):1851–1853
Burrell A, Foy C, Burns M (2011) Applicability of three alternative instruments for food authenticity analysis: GMO identification. Biotechnol Res Int 2011:838232. doi:10.4061/2011/838232
Fleige S, Pfaffl MW (2006) RNA integrity and the effect on the real-time qRT-PCR performance. Mol Aspects Med 27(2–3):126–139. doi:10.1016/j.mam.2005.12.003
Simbolo M, Gottardi M, Corbo V, Fassan M, Mafficini A, Malpeli G, Lawlor RT, Scarpa A (2013) DNA qualification workflow for next generation sequencing of histopathological samples. PLoS One 8(6):e62692. doi:10.1371/journal.pone.0062692
Kroneis T, Gutstein-Abo L, Kofler K, Hartmann M, Hartmann P, Alunni-Fabbroni M, Walcher W, Dohr G, Petek E, Guetta E, Sedlmayr P (2010) Automatic retrieval of single microchimeric cells and verification of identity by on-chip multiplex PCR. J Cell Mol Med 14(4):954–969
Acknowledgement
This work was supported by the EU SAFE Network of Excellence (LSHB-CT-2004-503243, EU 6th Framework Package) and the County of Styria, Austria.
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Kroneis, T., El-Heliebi, A. (2015). Quality Control of Isothermal Amplified DNA Based on Short Tandem Repeat Analysis. In: Kroneis, T. (eds) Whole Genome Amplification. Methods in Molecular Biology, vol 1347. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2990-0_10
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DOI: https://doi.org/10.1007/978-1-4939-2990-0_10
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-2989-4
Online ISBN: 978-1-4939-2990-0
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