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BioTechniques Volume 55, 2013 - Issue 5
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Microsatellite Marker Discovery using Single Molecule Real-Time Circular Consensus Sequencing on the Pacific Biosciences RS

Markus A. Grohme1 Molecular Biotechnology and Functional Genomics,Technical University of Applied Sciences Wildau,Wildau,GermanyView further author information
,
Roberto Frias Soler2 Department of Biology, Institute of Pharmacy and Molecular Biotechnology,Heidelberg University,Heidelberg,GermanyView further author information
,
Michael Wink2 Department of Biology, Institute of Pharmacy and Molecular Biotechnology,Heidelberg University,Heidelberg,GermanyView further author information
&
Marcus Frohme1 Molecular Biotechnology and Functional Genomics,Technical University of Applied Sciences Wildau,Wildau,GermanyCorrespondence[email protected]
View further author information
Pages 253-256 | Received 05 Jul 2013, Accepted 18 Oct 2013, Published online: 03 Apr 2018

Figures & data

Figure 1. Overview of microsatellite discovery using genomic shotgun circular consensus sequencing (CCS).

Genomic DNA is sheared to an appropriate size and hairpin adaptors are ligated to generate SMRTbell templates. During sequencing, a strand-displacing polymerase extends the sequencing primer (orange) and synthesizes the complementary strand in forward and reverse orientation. The resulting consensus sequences are processed using standard microsatellite identification tools.

Figure 1. Overview of microsatellite discovery using genomic shotgun circular consensus sequencing (CCS).Genomic DNA is sheared to an appropriate size and hairpin adaptors are ligated to generate SMRTbell templates. During sequencing, a strand-displacing polymerase extends the sequencing primer (orange) and synthesizes the complementary strand in forward and reverse orientation. The resulting consensus sequences are processed using standard microsatellite identification tools.
Figure 2. Average Phred quality scores of circular consensus sequences (CCSs).

The majority of CCS reads showed an average error probability between 1% and 0.1% (Phred score between 20 and 30).

Figure 2. Average Phred quality scores of circular consensus sequences (CCSs).The majority of CCS reads showed an average error probability between 1% and 0.1% (Phred score between 20 and 30).
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