Wednesday, September 5

1
20.1 1 Look at the illustration of Cloning a
Human Gene in a Bacterial Plasmid (Figure 20.4
in the orange book). If the medium used for
plating cells in step 5 did not contain
ampicillin, cells containing no plasmid would be
allowed to grow into colonies. What color would
those colonies be, and why?
2
Cloning a human gene in a bacterial plasmid
3
Cloning a human gene in a bacterial plasmid
Bacterial cell
Human cell
4
Isolate DNA
ampr gene
lacZ gene
Bacterial plasmid
Human chromosomes
5
Restriction enzymes
broken lacZ gene
ampr gene
gene of interest
Bacterial plasmid
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Mix together
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Mix together
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Add bacteria
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Add bacteria
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Grow on ampicillin
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Grow on ampicillin
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Add lactose mimic
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Add lactose mimic
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20.1 1 Look at the illustration of Cloning a
Human Gene in a Bacterial Plasmid (Figure 20.4
in the orange book). If the medium used for
plating cells in step 5 did not contain
ampicillin, cells containing no plasmid would be
allowed to grow into colonies. What color would
those colonies be, and why?
15
20.1 1 White (No functional lacZ gene is
present.)
16
20.1 2 Imagine you want to study human
ß-globin, a protein found in red blood cells. To
obtain sufficient amounts of the protein, you
decide to clone the ß-globin gene. Would you
construct a genomic library or a cDNA library?
What material would you use as a source of DNA or
RNA?
17
20.1 2 A cDNA library, made using mRNA from
developing red blood cells, which would be
expected to contain many copies of ß-globin
mRNAs.
18
20.1 3 What are two potential difficulties in
using plasmid vectors of human proteins from
cloned genes?
19
20.1 3 Some human genes are too large to be
incorporated into bacterial plasmids. Bacterial
cells lack the means to process RNA transcripts,
and even if the need for RNA processing is
avoided by using cDNA, bacteria lack enzymes to
catalyze the post-translational processing that
many human proteins undergo.
20
20.2 1 Suppose you carry out electrophoresis on
a sample of genomic DNA isolated from an
individual and treated with a restriction enzyme.
After staining the gel with a DNA-binding dye,
what would you see? Explain.
21
20.2 1 Any restriction enzyme will cut DNA in
many places, generating such a large number of
fragments that they would appear as a smear
rather than distinct bands when the gel is
stained after electrophoresis.
22
20.2 2 Explain why restriction fragment length
polymorphisms (RFLPs) can serve as genetic
markers even though they produce no visible
phenotypic differences.
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20.2 2 RFLPs are inherited in a Mendelian
fashion, and variations in RFLPs among
individuals can be detected by Southern blotting.
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20.3 1 What is a major difference between a
genetic (linkage) map and a physical map of a
chromosome?
25
20.3 1 In a genetic linkage map, genes and
other markers are ordered with respect to each
other, but only the relative distances between
them are known. In a physical map, the actual
distances between markers, expressed in base
pairs, are known.
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20.3 2 In general, how does the approach to
genome mapping used in the Human Genomic Project
differ from the shotgun approach?
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20.3 2 The three-stage approach employed in the
Human Genome Project involves genetic mapping,
physical mapping, and then sequencing of short,
overlapping fragments that previously have been
ordered relative to each other. continued
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20.3 2 The shotgun approach eliminates the
genetic mapping and physical mapping stages
instead, short fragments generated by multiple
restriction enzymes are sequenced and then
subsequently ordered by computer programs that
identify overlapping regions.
29
20.4 1 Current estimates are that the human
genome contains about 25,000 genes, but there is
evidence for many more different polypeptides.
What process might explain this discrepancy?
30
20.4 1 Alternative splicing of RNA transcripts
from a gene and post-translational processing of
polypeptides.
31
20.4 2 What is the major value of DNA
microarray analysis for studying gene expression?
32
20.4 2 It allows the expression of thousands of
genes to be expressed simultaneously, thus
providing a genome-wide view of which genes are
expressed in different tissues, under particular
conditions, or at different stages of development.
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20.4 3 Why is the genetic variation among
people so much less than it is among individuals
of many other species?
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20.4 3 Because the human species arose more
recently than many other species, there has been
less time for genetic variations in coding and
noncoding DNA to accumulate.
35
20.5 1 What is the advantage of using stem
cells for gene therapy?
36
20.5 1 Stem cells continue to reproduce
themselves.
37
20.5 2 List at least three different properties
that have been acquired by crop plants via
genetic engineering.
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• 20.5 2
• Herbicide resistance
• Pest resistance
• Disease resistance
• Delayed ripening
• Improved nutritional value