Quantitative and Qualitative Analyses of the SNRPN Gene Using Real-Time PCR with Melting Curve Analysis Chia-Cheng Hung, Shin-Yu Lin, Shuan-Pei Lin,

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Quantitative and Qualitative Analyses of the SNRPN Gene Using Real-Time PCR with Melting Curve Analysis Chia-Cheng Hung, Shin-Yu Lin, Shuan-Pei Lin, Chih-Ping Chen, Lang-Yao Chen, Chien-Nan Lee, Yi-Ning Su The Journal of Molecular Diagnostics Volume 13, Issue 6, Pages 609-613 (November 2011) DOI: 10.1016/j.jmoldx.2011.06.005 Copyright © 2011 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 1 Amplification curves of SYBR Green I-based real-time PCR analysis for detection of gene copy numbers of the SNRPN gene. The settings of the real-time PCR are the cycle range set between 22 and 34 cycles, the noise band set at 4.7863, and the threshold of estimation of the threshold cycle value was 9.2780. The results are shown by green (unaffected individuals), pink [Prader-Willi syndrome (PWS) with uniparental disomy (UPD) type], red (PWS with deletion type), and gray [Angelman syndrome (AS) with deletion type]. The Journal of Molecular Diagnostics 2011 13, 609-613DOI: (10.1016/j.jmoldx.2011.06.005) Copyright © 2011 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 2 The scatter plots of the calculated copy numbers by SYBR green-I–based real-time PCR are illustrated for the wild type, Prader-Willi syndrome (PWS) with deletion, PWS with uniparental disomy, and Angelman syndrome (AS) with deletion. UPD, uniparental disomy. The Journal of Molecular Diagnostics 2011 13, 609-613DOI: (10.1016/j.jmoldx.2011.06.005) Copyright © 2011 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 3 A: Normalized melting curve analysis of the SNRPN gene, and (B) the derivative plot [negative derivative of the fluorescence (F) with respect to temperature (T) versus temperature] of the SNRPN gene. The melting curves in blue are Prader-Willi syndrome (PWS) samples, the melting curves in red are wild types, and the melting curves in green are Angelman syndrome samples. The unmethylated melting peak was at 85°C, and the methylated melting peak was at 89°C. The results are shown by green (unaffected individuals), pink [PWS with uniparental disomy (UPD) type], red (PWS with deletion type), and gray [Angelman syndrome (AS) with deletion type]. The Journal of Molecular Diagnostics 2011 13, 609-613DOI: (10.1016/j.jmoldx.2011.06.005) Copyright © 2011 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions