© 2013 Elsevier, Inc. All rights reserved. Chapter 10 Author: Kelly Elkins © 2013 Elsevier, Inc. All rights reserved.

© 2013 Elsevier, Inc. All rights reserved. Figure 10.1 PCR primer locations for TPOX primers used to amplify Homo sapiens thyroid peroxidase (TPO) on chromosome 2; NCBI Nucleotide Sequence (Accession M68651): repeat (gray), and sample designed primers (arrows and bold). © 2013 Elsevier, Inc. All rights reserved.

© 2013 Elsevier, Inc. All rights reserved. Figure 10.2 Exponential amplification curves that show the results of forty cycles of gradient real-time PCR using primers depicted in Figure 10.1. Curves of amplified K562 DNA (from 1 ng starting material) as detected by SYBR green I. © 2013 Elsevier, Inc. All rights reserved.

© 2013 Elsevier, Inc. All rights reserved. Figure 10.3 A. Melting curve plot from 50-95 °C showing the temperature inflection at 79.5 °C for the K562 DNA (left image). B. First derivative plot of the melting curve from 50-95 °C showing the predominant peak at 79.5 °C for the K562 DNA (right image). © 2013 Elsevier, Inc. All rights reserved.

© 2013 Elsevier, Inc. All rights reserved. Figure 10.4 Sample 2% agarose gel of PCR of K562 DNA using designed primers. Lanes 1 & 7 contain the DNA Logic Ladder (sizes indicated), Lane 2 contains a low molecular weight ladder (500 - 20 bp) (sizes indicated) and Lanes 3-6 and 8-9 contain amplicons produced using experimental designed PCR primers. Lane 5 contains the amplicon produced from the primers shown in Figure 10.1. © 2013 Elsevier, Inc. All rights reserved.

© 2013 Elsevier, Inc. All rights reserved. Figure 10.5 Setup of gradient PCR amplification cycle and plate temperature protocol on BioRad iQ5. © 2013 Elsevier, Inc. All rights reserved.

© 2013 Elsevier, Inc. All rights reserved. Figure 10.6 Setup of gradient PCR run protocol on BioRad iQ5 © 2013 Elsevier, Inc. All rights reserved.