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    Ch.3 Dna Technology

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    วรรณศิริ คงสมนึก
    The document discusses several techniques used in DNA technology and genetic engineering. It describes (1) cutting and pasting DNA using restriction enzymes to insert a gene into a plasmid, (2) transforming bacteria by introducing plasmids containing the gene, and (3) selecting bacteria that contain the plasmid using antibiotics. It also explains polymerase chain reaction (PCR) to amplify a specific DNA region, gel electrophoresis to separate DNA fragments by size, and Sanger sequencing to determine the DNA nucleotide sequence.

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    Ch.3 Dna Technology

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    วรรณศิริ คงสมนึก
    13 views35 pages
    The document discusses several techniques used in DNA technology and genetic engineering. It describes (1) cutting and pasting DNA using restriction enzymes to insert a gene into a plasmid, (2) transforming bacteria by introducing plasmids containing the gene, and (3) selecting bacteria that contain the plasmid using antibiotics. It also explains polymerase chain reaction (PCR) to amplify a specific DNA region, gel electrophoresis to separate DNA fragments by size, and Sanger sequencing to determine the DNA nucleotide sequence.

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    The document discusses several techniques used in DNA technology and genetic engineering. It describes (1) cutting and pasting DNA using restriction enzymes to insert a gene into a plasmid, (2) transforming bacteria by introducing plasmids containing the gene, and (3) selecting bacteria that contain the plasmid using antibiotics. It also explains polymerase chain reaction (PCR) to amplify a specific DNA region, gel electrophoresis to separate DNA fragments by size, and Sanger sequencing to determine the DNA nucleotide sequence.

    Copyright:

    © All Rights Reserved
    Download as PDF, TXT or read online from Scribd
    Download as pdf or txt
    13 views35 pages

    Ch.3 Dna Technology

    Uploaded by

    วรรณศิริ คงสมนึก
    The document discusses several techniques used in DNA technology and genetic engineering. It describes (1) cutting and pasting DNA using restriction enzymes to insert a gene into a plasmid, (2) transforming bacteria by introducing plasmids containing the gene, and (3) selecting bacteria that contain the plasmid using antibiotics. It also explains polymerase chain reaction (PCR) to amplify a specific DNA region, gel electrophoresis to separate DNA fragments by size, and Sanger sequencing to determine the DNA nucleotide sequence.

    Copyright:

    © All Rights Reserved
    Download as PDF, TXT or read online from Scribd
    Download as pdf or txt
    of 35

    DNA

    TECHNOLOGY
    SUCHANUN SRIKAEW
    GENETIC
    ENGINEERING
    1) CUTTING & PASTING DNA
    • Use restriction enzymes cut out the gene
    of interest
    RESTRICTION ENZYMES
    RESTRICTION ENZYMES

    • blunt ends

    • sticky ends
    RESTRICTION ENZYMES
    1) CUTTING & PASTING DNA
    • Separately digest (cut) the gene fragment
    and the plasmid with EcoRI
    1) CUTTING & PASTING DNA
    • Separately digest (cut) the gene fragment
    and the plasmid with EcoRI
    1) CUTTING & PASTING DNA

    • Then, combine the


    fragments with DNA ligase
    RECOMBINANT DNA
    • The technology used for making artificial
    DNA by combining different DNA from
    different sources
    PLASMID

    • Origin of replication
    • Multiple cloning site
    • Selectable marker gene
    2) BACTERIAL TRANSFORMATION
    • Plasmids and other DNA can be introduced
    into bacteria
    3) BACTERIAL SELECTION
    • Use antibiotic selection and DNA analysis
    methods to identify bacteria that contain
    the plasmid
    POLYMERASE
    CHAIN
    REACTION

    • The technique to make


    many copies of a specific
    DNA region in vitro
    1) DENATURATION
    • (96°C): Heat the reaction strongly to separate,
    or denature, the DNA strands
    2) ANNEALING
    • (55 - 65°C): Cool the reaction so the primers
    can bind to their complementary sequences
    3) EXTENSION

    • (72°C): Raise the


    reaction temperatures
    so Taq polymerase
    extends the primers,
    synthesizing new
    strands of DNA
    POLYMERASE CHAIN REACTION
    • The cycle repeats 25-35 times which
    generally takes 2-4 hours, depending on
    the length of the DNA region being copied
    GEL
    ELECTROPHORESIS
    GEL ELECTROPHORESIS

    • The technique used


    to separate DNA
    fragments according
    to their size
    AGAROSE GEL
    • Gels for DNA separation are often made
    out of a polysaccharide
    GEL ELECTROPHORESIS
    • DNA samples are loaded into wells at one
    end of a gel, and an electric current is
    applied to pull them through the gel
    GEL ELECTROPHORESIS
    • When a gel is stained with a DNA-binding dye,
    the DNA fragments can be seen as bands

    • Ethidium
    bromide
    RFLP ANALYSIS
    HUMAN GENOME
    DNA SEQUENCING
    • The process of determining the sequence
    of nucleotide bases (As, Ts, Cs, and Gs) in
    a piece of DNA
    SANGER SEQUENCING

    • The target DNA


    is copied many
    times, making
    fragments of
    different lengths
    SANGER SEQUENCING
    • Fluorescent nucleotides mark the ends of
    the fragments and allow the sequence to
    be determined
    SANGER SEQUENCING
    NEXT-GENERATION SEQUENCING
    • new, large-scale approaches that increase
    the speed and reduce the cost of DNA
    sequencing

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