SeqSharp The Journal of Molecular Diagnostics

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SeqSharp The Journal of Molecular Diagnostics Dhruba J. SenGupta, Brad T. Cookson The Journal of Molecular Diagnostics Volume 12, Issue 3, Pages 272-277 (May 2010) DOI: 10.2353/jmoldx.2010.090134 Copyright © 2010 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 1 SeqSharp improves sequence quality from a two-step sequencing reaction. A: Schematic comparison of chain termination products from a two-step sequencing reaction and predicted electropherogram without (upper panel) and with SeqSharp (lower panel). B: Comparison of actual electropherograms obtained for S. aureus 16S rRNA gene sequence without (upper panel) or with SeqSharp (lower panel). The region of 16S sequence shown is between positions 239 and 271 within a 530-bp amplicon. Black arrowheads highlight a stretch of four bases where basecalls were affected within the section of electropherogram shown. Blue bars on top of bases indicate QV > 20. The Journal of Molecular Diagnostics 2010 12, 272-277DOI: (10.2353/jmoldx.2010.090134) Copyright © 2010 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 2 SeqSharp improves sequence quality from a one-step sequencing reaction. A: Schematic comparison of chain termination products from a one-step sequencing reaction and predicted electropherogram without (upper panel) and with SeqSharp (lower panel). B: Comparison of actual electropherograms obtained for spa sequence without (upper panel) or with SeqSharp (lower panel). The region of spa sequence shown is between positions 184 and 214 within a 367-bp amplicon. Black arrowheads highlight base positions where basecalls were wrong. C: One-step SeqSharp of N. farcinica 16S rRNA gene using dNTPs without dITP (upper panel) and with dITP (lower panel). The region of 16S sequence shown is between positions 354 and 381 within a 500-bp amplicon. Incorrect basecalls are highlighted by Black arrowheads. Blue bars on top of bases indicate QV > 20. The Journal of Molecular Diagnostics 2010 12, 272-277DOI: (10.2353/jmoldx.2010.090134) Copyright © 2010 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 3 One-step sequencing by SeqSharp using equimolar primer concentrations of forward and reverse primers. A: Complimentary strand sequences between positions 319 and 362 within a 500-bp amplicon from the N. farcinica 16S rRNA gene. B: Complimentary strand sequences between positions 360 and 405 within a 612-bp amplicon from the C. famata 26S rRNA gene. C: Complimentary strand sequences between positions 174 and 222 within a 310-bp amplicon from the human β-globin gene. Upper and lower panels show forward and reverse strands, respectively. Blue bars on top of bases indicate QV > 20. The Journal of Molecular Diagnostics 2010 12, 272-277DOI: (10.2353/jmoldx.2010.090134) Copyright © 2010 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions