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Chapter 12
DNA Technology and Genomics
(aka GENETIC ENGINEERING)
ALIGNED WITH
“Ch. 12 DNA Technology and Genomics Questions” Worksheet
1. What makes recombinant DNA
technology different from just DNA
technology? (12.1)
DNA Technology - method for studying &
manipulating genetic material
 Recombinant DNA – DNA in which two
different sources are combined into the
same DNA molecule

1. What are some examples of
recombinant DNA technology?
(12.1)
Mass production of materials
(cancer drugs, pesticides, insulin)
 Transfer of genes from one organism
to another


Production of pest-resistant plants,
heartier crops, more muscular farm
animals
Piggies + Jellyfish Gene
While these glowing pigs may be just “fun,” Asian
researchers have created a glowing fish whose
luminescence can be used as an indicator of the
presence or absence of certain chemical pollutants
2. What are plasmids? What makes them
such a good tool in gene cloning? (12.1)

Plasmid – small, circular DNA
not part of chromosome

Plasmids can carry foreign
genes (genes from other
organisms) and replicate
QUICKLY and CHEAPLY in the
lab  good for making LOTS of
copies of the foreign gene AND
the protein product of the gene
in mass quantities
3. Briefly describe the process of
gene cloning. (12.2)



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Bacterial plasmid (vector) and gene of
interest (already cut by RE) are mixed…
complimentary base pairing assures that
GOI (gene of interest) gets incorporated
into plasmid (step 3)
DNA ligase covalently bonds nucleotide
backbone (step 4)
Result is a recombinant plasmid which,
when inserted into a bacterial cell, will
multiply the new DNA (clone) (steps 5-6)
Note: the plasmid vector usually also
contains an antibiotic resistance gene that
will allow scientists to isolate colonies that
have the GOI. (Will grow bacteria on
pates w/antibiotic – those with out the
plasmid will be killed, those with plasmid
can survive).
4. What is genetic engineering and what
is its role in biotechnology? (12.1)

direct manipulation of genes for
practical purposes


ex: DNA fingerprinting, genetically
modified organisms and food,
transplantation of genes, cloning
Biotechnology uses GE to use
organisms and their components to
make useful products
5. What are restriction enzymes and how
do they function in cloning genes? (12.2)




Restriction Enzyme (RE) – molecular
scissors which identify and “cut”
certain nucleotide sequences,
creating restriction fragments. Leave
either blunt ends (not useful) or
“sticky ends” which can be combined
with other sticky ends (see next slide)
In nature, these enzymes protect
bacterial cells from “intruder” DNA
from bacteriophages (will chop up
foreign DNA); bacteria protect their
own DNA from RE’s by chemical
modification (methylation – adding
methyl groups to their A’s and C’s)
Use in gene cloning: Isolate the gene
of interest using a restriction enzyme
Cut the donor DNA & plasmid with the
same RE to make complimentary
sticky ends!!
6. What is meant by the term “sticky ends”? Why are they so
important in genetic recombination? (12.2)
7. Why is it important to cut the two DNA sources with the
same restriction enzyme? (12.2)
EcoRI – cleaves GAATTC



Sticky ends: the single
stranded ends of
double stranded DNA
fragments; the
unpaired nucleotides
can pair with
complimentary bases
from another molecule
cut with the same
RE!!!!
Only sticky ends cut
with same RE will be
complimentary
http://highered.mcgrawhill.com/olc/dl/120078/bio37.swf
Sample RE’s and their
restriction sites
8. What is a genomic library? What
two types of vectors can be used to
create these libraries? (12.4)



When cutting with a RE, you
get not only the gene of
interest, but all possible
fragments resulting from that
cut!!
Genomic Library – the entire
collection of all the cloned DNA
fragments from a genome
Bacterial plasmids and
bacteriophages can be used
as vectors to create libraries
9. What is cDNA? Explain how reverse
transcriptase works to create cDNA. (12.5)


cDNA – DNA that is
complimentary to
processed mRNA;
obtained by
REVERSING
TRANSCRIPTION
from a mRNA
sequence (catalyzed
by reverse
transcriptase)
Single-stranded DNA
molecule then creates
a compliment using
DNA polymerase
cDNA animation
10. What is the advantage for using
cDNA in the cloning process? (12.5)


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The resulting cDNA lacks all the
introns…it only has the
important genetic information
because all the introns were
spliced out during RNA
processing
Since bacteria don’t have
introns, they can’t remove them
from a foreign DNA insert when
making the mRNA.
cDNA is used to clone human
genes
This technique also helps us to
see what part of the original
gene is intron and what is exon.
We can now compare original
gene (introns and exons) vs. the
cDNA (all exons)
Also useful in studying particular
cell functions
11. Describe the advantages for using bacteria,
yeast, and mammalian cells for mass production of
gene products. Give one example and its use for
each. (12.6)



1982: Humulin
The first recombinant drug made by bacteria and
approved by the FDA
BACTERIA
 Readily available & cheap
 Have plasmids
 Grow rapidly
 Produce large quantities of
gene and protein QUICKLY
YEAST
 Easy to grow
 Good at synthesizing eukaryotic
proteins (can remove introns)
 Have plasmids
MAMMALIAN CELLS
 Attach sugars correctly to
make glycoproteins
 Can mass produce gene
product by whole animals rather
than cells (sheep produce CF
treatment in milk)
12. Briefly describe how DNA technology is
being used to create vaccines. (12.7)

Vaccine – a harmless variant or derivative of a
pathogen (usually bacteria or virus) that is used to
prevent an infectious disease by stimulating an
immune response in the vaccine recipient (human)
 Genetically engineered cells can be used to
produce large amounts of the virus’ outer protein
coat (hepatitis B vaccine) HepB vaccine animation
 Mutant viruses can be made by altering one or
more of the genes; immune response still triggered
 Replace some genes in harmless virus to provide
immunity to several diseases simultaneously
13. Explain how DNA probes tag a gene.
(12.8)


A radioactively labeled
complement (the probe)
to the gene of interest is
created and introduced
to single stranded DNA
from a phage clone
Can be used to screen
an entire bacterial
colony to discover which
one(s) contain genes of
interest
Ti plasmid animation
GENETICALLY MODIFIED (GM) ORGANISMS

Use Ti plasmid of Agrobacterium tumefaciens as the
vector  GM plant
 ex: soybeans and cotton crops receive bacterial
genes to make them resistant to herbicides and
pests
 ex: “golden rice” = rice with a few daffodil
genes added. Rice plant can now make
B-carotene, needed for vitamin A production.
Vitamin A deficiency (and resulting blindness) is a
serious problem for HALF of the world who
depend on rice as their staple food.