Small Molecule Real Time Sequencing

This presentation gives an overview on how how the Next Generational Sequencing method of Small Molecule Real Time Sequencing works.

1. Next Generation Sequencing Single Molecule Real Time Method

2. Overview Uses actual DNA polymerase to sequence Enzymatic speed fluorescent probe not to the nucleotide

3. Step 1 A complex of a DNA template and active polymerase is immobilized at the bottom of the ZMW Image found at http://esciencecentral.org/ebooks/applications-of-molecular-genetics/dna-sequencing-sanger-and-nextgeneration-sequencing.php

4. Step 2 Phospho -linked nucleotides are introduced into the chamber of the ZMW. Image found at http://esciencecentral.org/ebooks/applications-of-molecular-genetics/dna-sequencing-sanger-and-nextgeneration-sequencing.php Image found at http://www.pacificbiosciences.com/products/smrt-technology /

5. Step 3 Each of the four nucleotides is labeled with a different colored fluorophore and loaded. Image found at http://esciencecentral.org/ebooks/applications-of-molecular-genetics/dna-sequencing-sanger-and-nextgeneration-sequencing.php Image found at http://www.umich.edu/~caparray/products/ngs/pacbio

6. Step 4  As a base is held in the detection volume, a light pulse is produced. Image found at http://esciencecentral.org/ebooks/applications-of-molecular-genetics/dna-sequencing-sanger-and-nextgeneration-sequencing.php

7. Step 5 Nucleotides held by the polymerase prior to incorporation emit an extended signal that identifies the base being incorporated. Image found at http ://www.omixon.com/the-basics-of-next-generation-sequencing / Image found at http://esciencecentral.org/ebooks/applications-of-molecular-genetics/dna-sequencing-sanger-and-nextgeneration-sequencing.php

8. Step 6  This process occurs in parallel in up to thousands of ZMWs that make up the SMRT cell. Image found at http://esciencecentral.org/ebooks/applications-of-molecular-genetics/dna-sequencing-sanger-and-nextgeneration-sequencing.php

9. Work Cited Ambry Genetics. Making Sense of NextGent Sequencing. Kelly Gonzalez, MS, CGC, and Senior Manager of Clinical Genomics. Available at http://www.ambrygen.com/sites/default/files/pdfs/NERRG_4-10-12_Making_Sense_of_NetGen_Sequencing_KG(3).pdf . Access verified May 21, 2014. Omixon . Allen Van Deynze , 2010; Solanaceae Coordinated Agricultural Project. Next Generation Sequencing. Available at http://www.omixon.com/the-basics-of-next-generation-sequencing / . Access verified May 21, 2014. DNA Sequencing, Sanger and Next-Generation Sequencing. Omics eBooks Group. (2014). Yao Fei , Genome Institute of Singapore. Available at http :// esciencecentral.org/ebooks/applications-of-molecular-genetics/dna-sequencing-sanger-and-nextgeneration-sequencing.php Access verified Aug 5, 2014. SMRT TECHNOLOGY . Pacific Biosciences. (2014). Available at http://www.pacificbiosciences.com/products/smrt-technology / Access verified Aug 5, 2014. PacBio RS II Sequencing. Microarray Core Facility. (2012). Available at http ://www.umich.edu/~caparray/products/ngs/pacbio / Access verified Aug 5, 2014 . DNA SEQUENCING/GENOMICS: Toward personalized medicine: 3G DNA sequencing. (March 2010). Stephen Turner. BioOptics . Available at http:// www.bioopticsworld.com/articles/print/volume-3/issue-3/features/dna-sequencing-genomics.html Access verified Aug 5, 2014.

10. Recommended Videos SMRT TECHNOLOGY http ://www.pacificbiosciences.com/products/smrt-technology / Introduction to SMRT Sequencing http://article.wn.com/view/2014/05/14/Single_Molecule_RealTime_SMRT_DNA_Sequencing_Technology_From_g /

11. Next Generation Sequencing Ion Torrent Method