Biology 423 Research Paper:

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Biology 423 Research Paper Genetics behind
cloning of a human gene
1 - 2 page outlines due Nov. 6
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Writing a scientific paper Choose a topic Pick
a genetic disease for which the responsible gene
has been cloned. You can find lists of these
diseases at the following Yahoo site
http//dir.yahoo.com/Health/Diseases_and_Conditio
ns/Genetic_Disorders/ The OMIM database is also a
useful place to get information to start.
Please do not choose Cystic Fibrosis or
Fibrodysplasia ossificans progressiva. Collect
papers Look for genetics studies, family
studies as well as physiology, biochemistry and
cell biology
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Make a title active statements work well eg.
Grants disease is caused by a defective
potassium pump Make an outline Define a title
for each section even each paragraph. Use
active statements again. eg. Grants disease is
a genetic disease that affects breathing The
Grants disease mutation is on Chromosome 7 A
mutation in a potassium pump is linked to Grants
disease (The two above will be the main
sections) Expression of wild type potassium
pump reverts Grants disease effects in
cultured cells Grants potassium pump
transcripts are expressed in lung
cells Treatment of patients with potassium pump
blockers has no effect on progress of disease.
Microarray analysis suggests potential
treatment. These can be used as section titles
or as topic sentences (see below)
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Choose figures and make them. You can copy
figures from some of the papers you read. If so,
cite them properly eg (from Smith et al., 1987).
The figure with its figure legend should be
comprehensible without reading the text. Write
paragraphs First sentence is a topic sentence
Last sentence is a concluding sentence active
voice dont try to sound academic define your
terms, (I am not a medical doctor) if referring
to a figure, define the figure but do not
duplicate the figure legend. Maintain the same
tense, either past or present. Make an abstract
to put at the beginning of the paper summarizing
the experiments you will describe, the results
and the conclusions.
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Citations When you discuss published work, cite
the paper. Do the citation in the first sentence
in which the study is mentioned. Eg. Seven large
families with a high incidence of cystic fibrosis
were surveyed for DNA markers linked to the
disease (Smith et al., 1987). References at the
end of the section in alphabetical order Smith
J, Jones, P.A. and White, K. 1987 Family studies
map cystic fibrosis to Chromosome 7 Genetics
130 147-156. Use the journal Cell as an
example of how to format the paper, the
citations and the references. There are several
nice reference managers available. We use
EndNote for making bibliographies and storing
references.
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Mapping genes by recombination frequency
Test cross to monitor recombination between
different genes Frequency of recombination is
directly related to distance between genes
(loci) on chromosome Three point cross
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Drosophila, a model organism for genetics
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Traits for our three point cross Body color
yellow vs wild type Bristles forked vs straight
(wild type) Crossveins crossveinless vs wild type
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Fig. 5.12
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Test cross
vg b pr / vg b r X vg b pr / vg b pr
Punnet square Male Female vgbpr vg bpr
vgb pr vg b pr vgb pr vg bpr vgb
pr vg b pr Vg b pr vgbpr vg bpr vgb
pr vg b pr vgb pr vg bpr vgb pr vg b
pr
11111111 ratio of phenotypes if genes are
not linked
If genes are linked, parental combinations of
alleles are overrepresented in progeny
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Fig. 5.12
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3 genes, which is in the middle?
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Fig. 5.13
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Calculate distance between pairs vg to pr add
up all classes with a recombination event
between vg and pr or vg and pr
252 241 13 9 525
Divide by total number of chromosomes scored
525/4197 X 100 cM 12.5
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Calculate distance between pairs pr to b add
up all classes with a recombination event
between pr and b or pr and b
131 118 13 9 271
Divide by total number of chromosomes scored
271/4197 X 100 cM 6.4
The distance between vg and b is the sum of the
distance between vg-pr and pr-b
12.3 6.4 17.7
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Fig. 5.12
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Fig. 5.15
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How do we map genes in humans?
Relative association of markers Allelic variants
will co-segregate if the genes are closely
linked on a chromosome. Map distances depend on
frequency of recombination
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To map a human genetic trait Look for
association between mapped markers and a trait
of interest
Markers can be traits, proteins or DNA
sequences Anything that is polymorphic can be
mapped
We can translate map position into DNA sequence
by determining the linkage between DNA-based
markers and traits.
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Pedigree Analysis symbols
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Screen family members for DNA markers linked to
trait
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Screen family members for RFLP markers linked to
trait
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RFLP polymorphisms reveal genetic differences
2. Separate DNA fragments by size on an agarose
gel 3. Hybridize to single copy radioactive
probe- Southern Blot
1. Cut genomic DNA with Restriction enzymes
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HindIII polymorphism is closely linked to disease
N D D N N D
D N N N
Marker G8 from a randomly chosen phage clone with
a 17.6 kb human DNA insert.
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Test degree of linkage odds of linkage
Data looks like M1 is linked to SF. Mother has
two M1 alleles. Her chromosome is uninformative,
like a test-cross. Father has two different M
alleles. Recombination of his alleles can be
seen in this pedigree. In this family, there
are 8 informative chromosomes. 1 has a
recombination event. therefore, a rough estimate
of map distance is 1/8X 100 cM 12.5
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Odds of Linkage is (Probability gene and marker
are linked at a certain map distance) divided
by (Probability they are unlinked). Maximum
likelihood odds of linkage Change estimated
linkage distance (?) to get the best LOD score
for the data. example In our case best odds
of linkage value for this pedigree is L(.10)
6.3 Log of L or LOD 0.8 Maximum LOD score
Zmax 0.8
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LOD score is used to determine if two traits are
linked in human pedigrees
LOD nomenclature

• ? recombinant fraction
• L(?) odds of linkage at 0 gt ? lt 0.5
• L(0.5) odds of independent assortment
• LogL(?)/L(0.5) log-of-odds ratio LOD Z
• LOD scores gt 3 indicate linkage
• LOD scores lt -2 indicate non-linkage

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Example linkage of marker and trait
Zmax maximum likelihood score (MLS)
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To achieve significant LOD score Combine odds
of linkage for many families p1(L)/p1(NL) x
p2(L)/p2(NL) xp3(L)/p3(NL) In practice we
combine the log of odds LOD1 LOD2
LOD3. Continue until LOD gt 3.0 before linkage is
accepted Linkage distance is based on the linkage
distance that gives the maximum value for the
data.
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If genes and markers are unlinked, the p(L)/p(NL)
will be lt1.0 in some families and the final LOD
score will be negative (lt0). Therefore, as you
add more families the LOD score will only
increase if the data of the majority of families
supports linkage.
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Summary
Genetic distance between two genes is measured by
the recombination frequency of alleles of those
genes. If genes are closely linked, alleles for
those traits will not become recombined as
often as if the genes are more distant on a
chromosome. Genome maps can be made using
combinations of data from relative
recombination distances of many loci. Human
genetic linkage between two traits or two genes
is measured using Log scores. Log scores
greater than 3.0 (1/1000) chance of non-linkage
are the minimum values accepted as evidence of
linkage in family studies.