Biotech - Sequencing of the human genome was completed by...

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Unformatted text preview: Sequencing of the human genome was completed by 2007 DNA sequencing has depended on advances in technology, starting with making recombinant DNA In recombinant DNA , nucleotide sequences from two different sources, often two species, are combined in vitro into the same DNA molecule Methods for making recombinant DNA are central to genetic engineering , the direct manipulation of genes for practical purposes DNA technology has revolutionized biotechnology , the manipulation of organisms or their genetic components to make useful products An example of DNA technology is the microarray, a measurement of gene expression of thousands of different genes 3 Vector DNA as a carrier for the DNA segment to be cloned When a vector is introduced into a living cell, it can replicate making many copies Common vectors are plasmid or viral Also need the gene of interest from chromosomal DNA digest DNA of chromosome Cell containing gene of interest Gene inserted into plasmid Plasmid put into bacterial cell Recombinant DNA (plasmid) Recombinant bacterium Bacterial chromosome Bacterium Gene of interest Host cell grown in culture to form a clone of cells containing the cloned gene of interest Plasmid Gene of Interest Protein expressed by gene of interest Basic research and various applications Copies of gene Protein harvested Basic research on gene Basic research on protein Gene for pest resistance inserted into plants Gene used to alter bacteria for cleaning up toxic waste Protein dissolves blood clots in heart attack therapy Human growth hor- mone treats stunted growth 2 4 1 3 Step 1 in gene cloning 4 Step 2 Insert chromosomal DNA into vector Cut DNA using restriction enzymes or restriction endonucleases Made naturally by bacteria as protection against bacteriophages Cuts at specific known restriction sites Most restriction sites palindromic May produce sticky ends DNA ligase must be used to permanently link DNA Result may be Recircularized vector with no gene of interest inserted Recombinant vector with gene of interest inserted Animation: Restriction Enzymes 5 Vector carries a selectable marker Presence of antibiotics selects for cells expressing amp R gene contains plasmid amp R gene codes for b-lactamase that degrades ampicillin, which normally kills bacteria 6 Step 3 Goal for recombinant vector to be taken up by bacteria Transformation plasmid vector Transfection viral vector After treatment, only cells with the plasmid will grow on plates treated with ampicillin 7 To eliminate recircularized vectors from further examination, lacZ gene part of vector Insertion of chromosomal DNA disrupts lacZ gene lacZ codes for b- galactosidase which cleaves colorless X-Gal into a blue dye Recircularized plasmids will form blue colonies Recombinant vectors will form white colonies Animation: Cloning a Gene 8 DNA library Treatment of chromosomal...
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This note was uploaded on 09/24/2009 for the course CHEM 333 taught by Professor Baird during the Spring '09 term at UC Davis.

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Biotech - Sequencing of the human genome was completed by...

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