BIOL309 Lecture Notes - Lecture 6: Massive Parallel Sequencing, Charge-Coupled Device, Dna Footprinting

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30 Oct 2017
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Principles of dna sequencing: 2 techniques: maxam-gilbert sequencing (only used with specialized techniques such as. New tech: fluorescently tagged nucleotides (detected by laser beam) Sources of errors runs of same nucleotide runs of g and c (causes dna polymerase to skip/miss/add an extra nucleotide: hairpin loops, can improve by sequencing several overlapping fragments. Dna is sheared, sometimes it is sized. Adapter is added (ogligonucleotide), sometimes on both end; adapters allow dna molecule to bind to a bead so there is only one molecule per bead. Then mixed with oil in presence of enzymes (taq polymerase, dntps) Pcr reaction goes on, millions of beads with one drop oil per bead to increase the signal of the sequencing detection. Bead are put in a well, surrounded by the enzymes that are needed to detect the addition of the nucleotides. Now we have template for dna synthesis, ccd camera can see the emission of light in any of the wells.

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