lec 21a - Lecture 21a: Molecular techniques DNA sequencing,...

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Lecture 21a: Molecular techniques DNA sequencing, PCR
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Separation of DNA fragments: gel electrophoresis DNA = negatively charged - + electric field migrates toward positive electrode
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Separation of DNA fragments: gel electrophoresis DNA fragments Ethidium bromide (or Crystal Violet) intercalates in the DNA -> increased fluorescence visualization with a UV light
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Separation of DNA fragments: gel electrophoresis Small DNA fragments (20-2000 bp) -> acrylamide gels Large DNA fragments (200-20000 bp) -> agarose gels Concentration of agarose/acrylamide -> size of the “pores” -> range of separation of DNA fragments 1% agarose 2% agarose
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Separation of DNA fragments: gel electrophoresis 1 st lane: „ladder‟ of DNA fragments of know sizes (-) (+)
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DNA sequencing Principle: synthesis of DNA fragments with labeled ends and 1 nucleotide length difference -> Use nucleotide analogue that incorporates but terminates DNA synthesis Dideoxyribonucleoside triphosphate Previously: radiolabeled Now: fluorescently labeled -> four colors
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lec 21a - Lecture 21a: Molecular techniques DNA sequencing,...

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