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Lec13notescompletepart2 - Bio1A Lecture 13 Part 2 Chapter...

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Bio1A Lecture 13 Part 2 1 Chapter 20 DNA Technology and Genomics Understanding and Manipulating Genomes Sequencing of the human genome was largely completed by 2003 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 DNA cloning permits production of multiple copies of a specific gene or other DNA segment To work directly with specific genes, scientists prepare gene-sized pieces of DNA in identical copies, a process called gene cloning Most methods for cloning pieces of DNA in the laboratory share general features, such as the use of bacteria and their plasmids Cloned genes are useful for making copies of a particular gene and producing a gene product Fig. 20-2 Using Restriction Enzymes to Make Recombinant DNA Bacterial restriction enzymes cut DNA molecules at DNA sequences called restriction sites A restriction enzyme usually makes many cuts, yielding restriction fragments The most useful restriction enzymes cut DNA in a staggered way, producing fragments with sticky ends ” that bond with complementary “sticky ends” of other fragments DNA ligase is an enzyme that seals the bonds between restriction fragments Fig. 20-3 Cloning a Eukaryotic Gene in a Bacterial Plasmid In gene cloning, the original plasmid is called a cloning vector A cloning vector is a DNA molecule that can carry foreign DNA into a cell and replicate there Producing Clones of Cells Cloning a human gene in a bacterial plasmid can be divided into six steps: 1. Vector and gene-source DNA are isolated 2. DNA is inserted into the vector 3. Human DNA fragments are mixed with cut plasmids, and base-pairing takes place 4. Recombinant plasmids are mixed with bacteria 5. The bacteria are plated and incubated 6. Cell clones with the right gene are identified Fig. 20-4_1-3
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Bio1A Lecture 13 Part 2 2 Identifying Clones Carrying a Gene of Interest A clone carrying the gene of interest can be identified with a nucleic acid probe having a sequence complementary to the gene This process is called nucleic acid hybridization An essential step in this process is denaturation of the cells’ DNA, separation of its two strands Fig. 20-5 Storing Cloned Genes in DNA Libraries A genomic library that is made using bacteria is the collection of recombinant vector clones produced by cloning DNA fragments from an entire genome A genomic library that is made using bacteriophages is stored as a collection of
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