Noncontinuously Binding Loop-Out Primers for Avoiding Problematic DNA Sequences in PCR and Sanger Sequencing Kelli Sumner, Jeffrey J. Swensen, Melinda.

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Noncontinuously Binding Loop-Out Primers for Avoiding Problematic DNA Sequences in PCR and Sanger Sequencing Kelli Sumner, Jeffrey J. Swensen, Melinda Procter, Mohamed Jama, Whitney Wooderchak-Donahue, Tracey Lewis, Michael Fong, Lindsey Hubley, Monica Schwarz, Youna Ha, Eleri Paul, Benjamin Brulotte, Elaine Lyon, Pinar Bayrak-Toydemir, Rong Mao, Genevieve Pont-Kingdon, D. Hunter Best The Journal of Molecular Diagnostics Volume 16, Issue 5, Pages 477-480 (September 2014) DOI: 10.1016/j.jmoldx.2014.04.005 Copyright © 2014 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 1 Sample containing the MEN1 rs509606 polymorphism (not shown); sequences from c.240_259. A: Amplification with a normal forward primer; the 4-bp deletion (black bar), c.249_252delGTCT, was undetectable. B: Amplification with a loop-out forward primer showed a clearly discernable 4-bp deletion, c.249_252delGTCT. The Journal of Molecular Diagnostics 2014 16, 477-480DOI: (10.1016/j.jmoldx.2014.04.005) Copyright © 2014 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 2 Amplification of ASS1 exon 9. Top: Amplification through the problematic region with a traditional primer. The sequencing signal decreased to zero shortly after the G-rich region. Bottom: Amplification with a loop-out forward primer removed the problematic region, resulting in consistent sequence signal throughout the read. The Journal of Molecular Diagnostics 2014 16, 477-480DOI: (10.1016/j.jmoldx.2014.04.005) Copyright © 2014 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions