Copyright Pearson Prentice Hall 13-2 Manipulating DNA Copyright Pearson Prentice Hall

The Tools of Molecular Biology How do scientists make changes to DNA? Copyright Pearson Prentice Hall

The Tools of Molecular Biology Scientists use several techniques to manipulate DNA. Chemicals, computers, and bacteria are used to work with DNA. Scientists use these tools in genetics research and biotechnology. Copyright Pearson Prentice Hall

The Tools of Molecular Biology In genetic engineering, biologists make changes in the DNA code of a living organism. Copyright Pearson Prentice Hall

The Tools of Molecular Biology DNA Extraction DNA can be extracted from most cells by a simple chemical procedure. The cells are opened and the DNA is separated from the other cell parts. Copyright Pearson Prentice Hall

The Tools of Molecular Biology Cutting DNA  Restriction enzymes act as “molecular scissors.” come from various types of bacteria allow scientists to more easily study and manipulate genes cut DNA at a specific nucleotide sequence called a restriction site Copyright Pearson Prentice Hall

The Tools of Molecular Biology Each restriction enzyme cuts DNA at a specific sequence of nucleotides. Molecular biologists have developed different techniques that allow them to study and change DNA molecules. Restriction enzymes cut DNA at specific sequences. This drawing shows how restriction enzymes are used to edit DNA. The restriction enzyme EcoR I, for example, finds the sequence CTTAAG on DNA. Then, the enzyme cuts the molecule at each occurrence of CTTAAG. The cut ends are called sticky ends because they may “stick” to complementary base sequences by means of hydrogen bonds. Copyright Pearson Prentice Hall

The Tools of Molecular Biology -some cut straight across and leave “blunt ends” - some make staggered cuts and leave “sticky ends” Molecular biologists have developed different techniques that allow them to study and change DNA molecules. Restriction enzymes cut DNA at specific sequences. This drawing shows how restriction enzymes are used to edit DNA. The restriction enzyme EcoR I, for example, finds the sequence CTTAAG on DNA. Then, the enzyme cuts the molecule at each occurrence of CTTAAG. The cut ends are called sticky ends because they may “stick” to complementary base sequences by means of hydrogen bonds. Copyright Pearson Prentice Hall 8

The Tools of Molecular Biology Separating DNA   In gel electrophoresis, DNA fragments are placed at one end of a porous gel, and an electric voltage is applied to the gel. The DNA fragments are separated based on size. Smaller fragments travel farther in a certain amount of time than larger fragments. Copyright Pearson Prentice Hall

The Tools of Molecular Biology DNA plus restriction enzyme Power source Longer fragments Shorter fragments Mixture of DNA fragments Gel Gel electrophoresis is used to separate DNA fragments. First, restriction enzymes cut DNA into fragments. The DNA fragments are then poured into wells on a gel, which is similar to a thick piece of gelatin. An electric voltage moves the DNA fragments across the gel. Because longer fragments of DNA move through the gel more slowly, they do not migrate as far across the gel as shorter fragments of DNA. Based on size, the DNA fragments make a pattern of bands on the gel. These bands can then be compared with other samples of DNA.  Gel Electrophoresis Copyright Pearson Prentice Hall

The Tools of Molecular Biology First, restriction enzymes cut DNA into fragments. The DNA fragments are poured into wells on a gel. DNA plus restriction enzyme Gel electrophoresis is used to separate DNA fragments. First, restriction enzymes cut DNA into fragments. The DNA fragments are then poured into wells on a gel, which is similar to a thick piece of gelatin. An electric voltage moves the DNA fragments across the gel. Because longer fragments of DNA move through the gel more slowly, they do not migrate as far across the gel as shorter fragments of DNA. Based on size, the DNA fragments make a pattern of bands on the gel. These bands can then be compared with other samples of DNA.  Mixture of DNA fragments Gel Gel Electrophoresis Copyright Pearson Prentice Hall

The Tools of Molecular Biology An electric voltage is applied to the gel. The smaller the DNA fragment, the faster and farther it will move across the gel. Power source Gel electrophoresis is used to separate DNA fragments. First, restriction enzymes cut DNA into fragments. The DNA fragments are then poured into wells on a gel, which is similar to a thick piece of gelatin. An electric voltage moves the DNA fragments across the gel. Because longer fragments of DNA move through the gel more slowly, they do not migrate as far across the gel as shorter fragments of DNA. Based on size, the DNA fragments make a pattern of bands on the gel. These bands can then be compared with other samples of DNA.  Copyright Pearson Prentice Hall

The Tools of Molecular Biology Power source Longer fragments Gel electrophoresis is used to separate DNA fragments. First, restriction enzymes cut DNA into fragments. The DNA fragments are then poured into wells on a gel, which is similar to a thick piece of gelatin. An electric voltage moves the DNA fragments across the gel. Because longer fragments of DNA move through the gel more slowly, they do not migrate as far across the gel as shorter fragments of DNA. Based on size, the DNA fragments make a pattern of bands on the gel. These bands can then be compared with other samples of DNA.  Copyright Pearson Prentice Hall

Copyright Pearson Prentice Hall Using the DNA Sequence Restriction Map A restriction map is a map of known restriction sites within a sequence of DNA that requires the use of restriction enzymes. Fragment sizes between restriction sites Does not show anything about genes or DNA sequence Copyright Pearson Prentice Hall

Copyright Pearson Prentice Hall Using the DNA Sequence Making Copies  Polymerase chain reaction (PCR) is a technique that quickly copies a desired segment of DNA. Small amounts of DNA can be multiplied making it easier to analyze. Made possible by an enzyme found in a bacterium living in hot springs in Yellow Stone National Park. Copyright Pearson Prentice Hall

Copyright Pearson Prentice Hall Using the DNA Sequence The three steps of PCR occur in a cycle. heat is used to separate double-stranded DNA molecules primers bind to each DNA strand on opposite ends of the segment to be copied DNA polymerase binds nucleotides together to form new strands of DNA DNA strands polymerase nucleotides primer 1 primer 2 Copyright Pearson Prentice Hall 16

Copyright Pearson Prentice Hall Using the DNA Sequence PCR amplifies DNA samples by doubling the number of DNA molecules. After repeating the steps 30 times, 1 billion copies of the original DNA can be made. Copyright Pearson Prentice Hall 17

New Horizons in Medicine with PCR Cancer warning-Scientists are developing tests to pick up the genetic changes which take place in cancer cells very early in the development of the disease Tissue matching-In organ transplants, a close tissue match between the donor and the recipient reduces the chance of rejection Forensic medicine-PCR, along with DNA fingerprinting, has provided major breakthroughs for identifying and eliminating suspects Genetic testing-PCR makes it easier to identify individuals who carry the genes which can cause problems like cystic fibrosis and muscular dystrophy Infection detection-can provide a speedy and accurate diagnosis for serious infections so treatment can be correctly given Copyright Pearson Prentice Hall

Copyright Pearson Prentice Hall 13-2 Copyright Pearson Prentice Hall

Copyright Pearson Prentice Hall 13-2 Restriction enzymes are used to extract DNA. cut DNA. separate DNA. replicate DNA. Copyright Pearson Prentice Hall

Copyright Pearson Prentice Hall 13-2 During gel electrophoresis, the smaller the DNA fragment is, the more slowly it moves. heavier it is. more quickly it moves. darker it stains. Copyright Pearson Prentice Hall

Copyright Pearson Prentice Hall 13-2 The DNA polymerase enzyme found in bacteria living in the hot springs of Yellowstone National Park illustrates genetic engineering. the importance of biodiversity to biotechnology. the polymerase chain reaction. selective breeding. Copyright Pearson Prentice Hall

Copyright Pearson Prentice Hall 13-2 A particular restriction enzyme is used to cut up DNA in random locations. cut DNA at a specific nucleotide sequence. extract DNA from cells. separate negatively charged DNA molecules. Copyright Pearson Prentice Hall

Copyright Pearson Prentice Hall 13-2 During gel electrophoresis, DNA fragments become separated because multiple copies of DNA are made. recombinant DNA is formed. DNA molecules are negatively charged. smaller DNA molecules move faster than larger fragments. Copyright Pearson Prentice Hall

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