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Reading Guide Answers for Chapter 10
10-1 What is Genetic Engineering?
In 1973 Cohen & Boyer constructed the first genetically engineered organism
A gene for r-RNA from a frog was inserted into the DNA of the bacterium Escherichia coli (E. coli)
The bacteria produced from r-RNA
This ushered in a scientific revolution in biology
A. The Basics of Genetic Engineering
Genetic Engineering = process used to isolate a gene from the DNA of one organism & transfer it into DNA of another organism
It involves building recombinant DNA
A molecule is made from pieces of DNA from separate organisms.
Steps in genetically engineering an organism
1. Cleaving DNA—cutting out the gene of interest
2. Producing recombinant DNA—DNA fragment is inserted into a vector
the agent used to carry the DNA to other organism
(1) viruses
(2) plasmids—circular DNA found in bacteria
3. Cloning cells—cells containing the inserted DNA are allowed to reproduce in large numbers
4. Screening cells—cells receiving gene are identified and isolated
To understand this process we will apply these steps to Cohen and Boyer’s Frog experiment
B. Cleaving DNA
Transferred a gene that codes for r-RNA
1. Used restriction enzymes to cut out the gene
a) Molecular scissors
b) Cuts DNA at specific sequences
c) Makes a lot of little segments of DNA that are unique
Have same nucleotides running in both directions (can be read either way and they are the same)
EcoR1 restriction enzyme cuts the sequence GAATTC
2. many restriction enzymes make a staggered cut and leaves “sticky ends”
a) they can pair with each other very conveniently
b) sealed with the enzyme ligase
c) can pair with another DNA fragment that has been cut with the same restriction enzyme
C. Producing Recombinant DNA
1. Cohen and Boyer used the restriction enzyme EcoR1
a) Cut the plasmid with this to use as the vector
b) Vector carried the r-RNA gene from the frog
2. isolated a gene fragment that had two important genes
a) gene for DNA replication
b) gene for resistance to antibiotic tetracycline
3. frog gene was inserted in plasmid then called recombinant DNA
D. Cloning Cells
1. The plasmids are then mixed with bacteria in the presence of solutions that aid in the uptake of the plasmids by the bacteria. Only some of the bacteria will take in the recombined plasmid.
2. Antibiotic resistant gene for tetracycline is now important
a) Tetracycline is added to the bacteria
b) Only the cells that were NOT killed were the ones that took up the vector (plasmid with frog DNA)
c) These cells were allowed to reproduce
Therefore the plasmid with the gene was cloned
E. Screening Cells
1. checked to see if the frog gene was present
2. Southern Blot—technique used to identify a gene
a) Uses radioactive RNA or DNA as a “probe”
b) Specifically ordered nucleotides complementary to gene to be identified
c) Cloned DNA is cut into fragments by restriction enzymes
d) DNA Fragments are separated by size using gel electrophoresis
(1) Fragments are made using restriction enzymes.
(2) DNA fragments placed in indentions (wells) at one end of a block of gel
(3) Electrical current is run through the gel
(4) DNA is negatively charged and is drawn towards the opposite end.
(5) Fragments move according to their size. Smaller fragments move faster than larger ones.
(6) When current is turned off, gel is stained and bands appear where each group of DNA fragments stopped.
F. Identifying Sequences of DNA
Tools of the biotechnologist
1. RFLP’s – restriction fragment length polymorphism
a) Restriction enzymes cut at specific places with specific lengths
b) Because differences in nucleotide sequences the fragments are different sizes
c) Different organisms have different fragment lengths
d) When separated by gel electrophoresis forms a
2. DNA fingerprint
a) Bands represent a unique arrangement of RFLP sites
b) They are unique to an individual; can be used to establish identity
c) Used on blood, semen, bone, and hair can be used
3. PCR—Polymerase Chain Reaction
a) Makes unlimited copies of DNA (a gene)
b) Uses DNA polymerase and supply of all four nucleotides
c) Revolutionized research in molecular biology
4. Mapping the Human Genome (this has been completed)
a) Identify and locate all genes in a human cell.
b) Goals of project
(1) Improve human genetic maps
(2) Construct physical maps of entire chromosomes
(3) Determine complete DNA sequence in human genome
(4) Future—develop new methods for mapping and sequencing DNA
10-2 The New Medicine
Curing and preventing illnesses
Proteins as medicines
New vaccines
Replacement of defective genes.
A. Making Genetically Engineered Drugs
Failure of critical proteins causes illnesses
1. Diabetes mellitus type I (insulin-dependent)
a) Body cannot make right amount of insulin
b) Treated by injecting insulin
c) Body normally produces it in small amounts
2. insulin genetically engineered by bacteria
first commercially produced item in 1982
3. anticoagulants—dissolves blood clots, helps in heart attacks
4. factor VIII—promotes blood clotting; safer than from real blood
B. Making genetically engineered vaccines
1. What are they?
a) Harmless version of pathogen
b) Immune system recognizes pathogens’ surface proteins
c) Immune system response is to make antibodies
d) Antibodies stop growth of pathogen
2. How are they made?
a) Killed pathogen or make it unable to grow
b) Failure of process may cause disease
c) Most are safe, but small danger
3. A newer, safer method uses genetic engineering
a) Uses genes of pathogens surface proteins
b) Inserted in DNA of harmless bacteria
c) Bacteria becomes a safe vaccine
4. genetically engineered vaccines
a) herpes II (sexually transmitted disease)
b) hepatitis B (liver disease)
c) goal to produce a malaria vaccine
C. Curing Genetic Disorders
Caused by non-functioning copy of a gene
Solution is to give person a working copy
1. problems
a) defective gene is difficult to identify & isolate
b) hard to transfer good copy into cell that need it
c) hard to keep altered cells alive for long
2. Gene Therapy
Replacing defective gene with a healthy gene and inserting it into bone marrow which will then mass produce the cells WITH the healthy gene
3. TNF (tumor necrosis factor)
a) Fights cancer
b) White blood cells secrete this protein
c) Attack and kill cancer cells
10-3 The New Agriculture
Crops more resistant to plant diseases & pests
Tomatoes with longer shelf-life
Milk production increased
Increased growth rate in livestock
A. Transporting Genes In to Plants
1. crown gall – plant tumor
a) bacterial plasmid
b) Ti plasmid (tumor- inducing)
2. infects tomatoes, tobacco and soybeans
3. plasmid inserts itself into plant cell’s chromosome
4. how is the vector made?
a) Tumor gene removed from plasmid
b) Vacancy filled with DNA of interest
c) DNA then carried to chromosome of plant
B. Making Crops Resistant to Herbicides & Insects
1. development of crops plants resistant to glyphosate
a) lowers cost of crop because does not need weeding
b) weeds die and crop is resistant so it thrives
c) benefits environment due to biodegradability
2. cotton plants resistant to pests so no chemicals needed to kill the insects or pests
C. Developing Crops That Need No Fertilizer
1. All plants need nitrogen to make proteins and DNA
2. most abundant source is air
(1) plants cannot use it
(2) must come from soil and roots
(3) farmers must add fertilizers rich in N to soil
3. idea is to produce plants that can fix nitrogen from air to use
D. Improving Livestock Production
1. growth hormone given to cows to improve milk production
2. bacteria are producing the hormone
3. added as a supplement to cow’s diet
4. growth hormone added to diet of cows and hogs to increase their weight
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