Frontiers of Biotechnology

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Frontiers of Biotechnology
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Manipulating DNA

• Biotechnology is used to identify people, produce
  transgenic organisms and clones, study diseases
  and evolution, and create medical treatments for
  people with life threatening diseases.

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Techniques to Manipulate DNA

• Scientists must be able to work with DNA without
  being able to see or handle it directly.
• Scientists use artificial nucleotides, artificial
  genes, chemical mutagens, computers, enzymes,
  bacteria, and many other techniques to manipulate
  DNA.

Spooled DNA
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Restriction Enzymes

• Restriction enzymes are enzymes that cut DNA
  molecules at specific nucleotide sequences.
• Any time the restriction enzyme comes across the
  specific nucleotide sequence, it cuts the DNA.
• This allows scientists to work with small pieces
  of DNA at a time.
• Restriction enzymes are produced naturally by
  bacteria to cut the DNA of invading viruses.
• Restriction enzymes can either make clean cuts
  (blunt ends) of the DNA or leave sticky ends
• The sticky ends are staggered cuts in the DNA,
  that allow the DNA to reform easily.

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Restriction Maps

• Gel Electrophoresis
• Once DNA has been cut by restriction enzymes, it
  is placed into a gel electrophoresis plate.
• Gel electrophoresis uses an electrical current to
  separate a mixture of DNA fragments from each
  other.
• A positive electrode is set at one end, and a
  negative electrode is set at the opposite end.
• Because DNA has a negative charge, the fragments
  move toward the positive end.
• The larger fragments move slower than the smaller
  fragments, therefore the length of the DNA
  fragment can be estimated by the distance it
  travels through the gel in a certain period of
  time.

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Restriction Maps

• Restriction maps use gel electrophoresis to show
  the lengths of DNA fragments between restriction
  sites in a strand of DNA.
• Restriction maps are used to study mutations.
• They will show if nucleotides have been added or
  deleted from a particular strand of DNA.
• Or, a mutation may lead to a restriction site,
  and the DNA would not be cut in the same place.

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Copying DNA

• Forensic scientists use DNA from cells in a
  single hair at a crime scene to identify a
  criminal.
• Doctors test a patients blood to quickly detect
  the presence of bacteria that causes Lyme
  disease.
• Scientists compare DNA from different species to
  determine how closely the species are related.
• If the original DNA from any of these sources is
  too small to accurately study, the samples of DNA
  must be increased, or amplified, so that they can
  be analyzed.

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Polymerase Chain Reaction

• PCR is a technique that produces millionsor even
  billionsof copies of a specific DNA sequence in
  just a few hours.
• It is a very simple process.
• There are four materials involved the DNA to be
  copied, DNA polymerases, large amounts of each of
  the four nucleotides (A, T, C, G), and two
  primers.
• A primer is a short segment of DNA that acts as
  the starting point for a new strand.
• Each PCR cycle doubles the number of DNA copies.
  The original piece of DNA becomes two copies.
  Those two copies become fourect.

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PCR Process

• 1. Separating.
• The container with all the reactants is heated to
  separate the double-stranded DNA into single
  strands.
• 2. Binding.
• The container is cooled and the primers bind to
  their complimentary DNA sequences. One primer
  binds to each DNA strand. The primers bind on the
  opposite ends of the DNA segment being copied.
• 3. Copying.
• The container is heated again and the polymerases
  begin to build new strands of DNA. Added
  nucleotides bind to the original DNA strands by
  complimentary base pairing. The polymerases
  continue attaching nucleotides until the entire
  DNA segment has been copied.

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DNA Fingerprinting

• DNA evidence is used to convict a criminal,
  release an innocent person from prison, or solve
  a mystery.
• A couple of decades ago, the lines and swirls of
  someones fingertip were a detectives best hope
  for identifying someone. Now, investigators
  gather biological samples and analyze DNA for
  another kind of evidence a DNA fingerprint.

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DNA Fingerprinting

• A DNA fingerprint is a kind of restriction map.
• It is a representation of parts of an
  individuals DNA that can be used to identify a
  person at a molecular level.
• People differ greatly in the number of repeated
  non-coding sequences of DNA.
• DNA fingerprints can also be used to show
  relationships between family members.
• The children have similar DNA fingerprints to
  each other, but they are not identical. Also,
  their DNA fingerprints are combinations of the
  DNA fingerprints of the parents.

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DNA Fingerprinting and Identification

• The reliability of DNA identification relies on
  probability.
• Ex. Suppose that 1 in every 500 people has three
  copies of the repeat at location A.
• That means that any person has a 1 in 500 chance
  of having a matching DNA fingerprint for that
  region of their chromosome.
• Then, suppose that 1 in 90 people has six copies
  of the repeat sequence at location B, and 1 in
  120 people has ten copies of the repeat sequence
  at location C.
• Individual probabilities are multiplied by each
  other to find total probability. Therefore, when
  the three separate probabilities are multiplied,
  suddenly the chance that two people have the same
  DNA fingerprint is very small.
• 1/500 x 1/90 x 1/120 1/5,400,000 1 chance in
  5.4 million people.

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Uses of DNA Fingerprinting

• DNA fingerprinting can be used to convict a
  criminal or set an innocent person free.
• The Innocence Project has successfully released
  249 wrongly convicted people from jail using DNA
  fingerprinting.
• DNA fingerprinting can be used to identify
  familial relationships (paternity).
• DNA fingerprinting is also used to study
  biodiversity in an area, identify genetically
  engineered crops, and to follow migration
  patterns of native species.

Larry Fuller spent 18 years in prison, after
being wrongfully convicted of aggravated rape in
1981. Fuller was excluded as the rapist after
advanced DNA testing. He was released in January
2007.
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Genetic Engineering

• Glowing mice are used in cancer research, glowing
  plants are used to track genetically modified
  crops, and in 1999, British researchers
  introduced glowing yeast cells.
• The glowing yeast cells can detect pollution in
  an environment.
• Under normal conditions, the cells do not glow,
  but when they come into contact with certain
  chemicals, they glow.
• This indicated areas that need to be cleaned up.

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Cloning

• A clone is a genetically identical copy of a gene
  or of an organism.
• Cloning can be a natural process.
• Plants can clone themselves, bacteria produce
  genetically identical copies of themselves, and
  identical twins are clones of each other.
• To clone a mammal, scientists swap DNA between
  cells with a technique called nuclear transfer.
• 1. An unfertilized egg is taken from an animal,
  and the eggs nucleus is removed.
• 2. The nucleus of a cell from the animal to be
  cloned is implanted into the egg.
• 3. The egg is stimulated, and if the transfer is
  successful, the egg will begin dividing.

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Dolly The First Clone

• Born July 5th, 1996 in Endinburgh, Scotlandwas
  the fist cloned mammal (cloned from a mammary
  gland, and named after Dolly Parton)
• She was the only lamb who survived to adulthood,
  out of 227 attempts.
• She gave birth to 6 lambs, and died at the age of
  6 due to lung disease.

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The Future of Cloning

• In January 2009, scientists in Spain successfully
  cloned a Pyrenean Ibexa species declared extinct
  in 2000.
• They used skin cells preserved in liquid
  nitrogen.
• The Ibex died shortly after birth due to physical
  defects in its lungs.
• The possibility of cloning other endangered or
  extinct species (like the wooly mammoth or
  dinosaurs) is closer.
• This still does not increase genetic diversity in
  breeding pools however, and does not help loss of
  habitat.

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Genetic Engineering

• Genetic research relies on cloning, but not of
  entire organisms.
• Instead, the cloning of individual genes is used
  to make a copy of one segment of DNA.
• In some cases, scientists insert cloned genes
  from one organism into an entirely different
  organism.
• Genetic engineering is when you change an
  organisms DNA to give an organism new traits.
• Genetic engineering is only possible because the
  genetic code is shared by all organisms.

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Recombinant DNA

• Recombinant DNA is DNA that contains genes from
  more than one organism.
• To create recombinant DNA, scientists often used
  plasmids located in bacteria.
• Plasmids are closed loops of DNA within the
  bacteria that are easily manipulated.
• Scientists are using recombinant DNA to
• Try to create crop plants that produce medicines
  and vitamins.
• Try to create vaccines against HIV.
• Make hormones like HGH, insulin, and oxytocin.
• Use in gene therapy.
• ETC!

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Transgenic Organisms

• A transgenic organism has one or more genes from
  another organism inserted into the genome.
• Ex. The gene for human insulin can be put into
  plasmids. The plasmids are inserted into
  bacteria. The transgenic bacteria make human
  insulin which can be collected and used to treat
  people with diabetes.

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Transgenic Plants

• Scientists create transgenic plants by inserting
  a gene into a bacterias plasmid and having the
  bacteria infect a plant. The infected plant will
  then incorporate the new gene into its DNA.
• This technique has allowed scientists to give
  plants new traits, such as resistance to frost,
  diseases, and insects.
• This increases crop yields, more food is produced
  more quickly and cheaply.
• Genetically modified (GM) foods, are now common
  in the United States.

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Transgenic Animals

• Transgenic animals are difficult to make. It take
  many trials (hundreds) before a transgenic animal
  will form correctly to adulthood.
• The advantage with transgenic animals is that the
  transgenic gene will be present in ALL of their
  DNA, including in their reproductive cell. So,
  transgenic traits will be passed on to the next
  generation.
• Transgenic mice are often used as models to study
  human development and disease.
• They are used to study cancer (oncomice),
  diabetes, brain function and development, sex
  determination, etc.
• Other mice are called gene knockout mice.
• These mice are used to student gene functions and
  point mutations.

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Concerns About Genetic Engineering

• There are concerns regarding human health and the
  environment.
• People routinely eat GM foods without knowing it.
  Scientists have not been able to discover any
  adverse health effects so far.
• Critics claim that not enough research has been
  done on possible allergic reactions or other
  unknown side effects.
• Another concern is that GM plants may cause bees
  and butterflies to go extinct (by transgenically
  producing pesticides).
• Transgenic plants may also cross-pollinate with
  wild natural plants.
• Finally, transgenic plants may decrease genetic
  diversity in crops and leave them more vulnerable
  to new diseases or pests.
• Another concern is about the ethics of genetic
  engineering in the first place.

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Genomics

• Genomics is the study of genomes, which can
  include the sequencing of all of an organisms
  DNA.
• This is how we know that humans and chimpanzees
  share 99-98 of their DNA.
• Comparing DNA from many people at one time helps
  researchers find genes that cause disease, and it
  helps them understand how medication works.
• Biologists study DNA of different species to
  learn how closely related they are to each other
  and to extrapolate how far back in the
  evolutionary time line they diverged.

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DNA Sequencing

• All genomic studies begin with DNA sequencing.
• This is determining the order of DNA nucleotides
  in genes or in genomes.
• PCR is one method.
• Humans do not have the largest genome. Vanilla
  plants, crested newts, and lungfish are among the
  many organisms who have a larger genome than us!

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The Human Genome Project

• There are 30,000-40,000 genes in the human
  genome.
• Each gene represents about 100,000 DNA bases.
• The Human Genome Project has two goals
• 1. To map and sequence all of the DNA base pairs
  of the human chromsomes.
• 2. To identify all of the genes within the
  sequence.
• Right now, the Human Genome Project is working on
  the HapMapthe study of how DNA sequences vary
  among people.
• This will hopefully identify genetic differences
  that play a part in human diseases.

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Bioinformatics

• Bioinformatics is the use of computer databases
  to organize and analyze biological data.
• Bioinformatics give scientists a way to store,
  share, and find data.

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Genetic Screening

• Genetic screening is the process of testing DNA
  to determine a persons risk of having or passing
  on a genetic disorder.
• It often involved both pedigree analysis and DNA
  tests.
• There are tests for about 900 genetic disorders,
  including cystic fibrosis, Duchennes muscular
  dystrophy, and breast cancer.

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Gene Therapy

• Gene therapy is the replacement of a defective or
  missing gene, or addition of a new gene, into a
  persons genome to treat a disease.
• Scientists will often use de-natured viruses to
  introduce the new gene to the body.
• There have been a few successful cases of gene
  therapy wiping out diseases.
• There are many experiments going on with gene
  therapy.
• Scientists are trying to insert genes into the
  immune system that stimulated the immune system
  to attack cancer cells.
• Another method is to insert suicide genes into
  cancer cells.