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Balanced Translocation detected by FISH

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Assign numbers to results of linkage analysis. to deal with non-ideal meioses ... F506. Normal. Testing for specific mutations. ARMS 3' mis-match of primer ... – PowerPoint PPT presentation

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Title: Balanced Translocation detected by FISH


1
Balanced Translocation detected by FISH
2
Red- Chrom. 5 probe
Green- Chrom. 8 probe
3
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4
2D Protein Gels
5
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6
MS-peptide size signature match to all predicted
proteins
7
POSITIONAL CLONING
8
X
2
3
Parent
X
Gamete
X
Child
9
A1
B1
C1
D1
X
2
3
E2
Parent
B2
C2
A2
A2
B1
C1
D2
X
Gamete
B1
C1
A2
X
Child
10
A1
B1
C1
D1
X
2
3
E2
Parent
B2
C2
A2
A2
B1
C1
D2
X
NR
Gamete
R
B1
C1
A2
X
Child
11
Positional Cloning by Recombination Mapping
  • Follow the mutation
  • 2. Follow which DNAs are
  • co-inherited (linked)

12
Positional Cloning by Recombination Mapping
  • Follow the mutation
  • To determine disease gene
  • presence or absence (genotype)
  • from phenotype you must
  • first establish
  • Dominant / recessive
  • Aurosomal / sex-linked

13
SINGLE GENE DEFECTS
Modes of Inheritance
To deduce who (likely) has one or two copies of
mutant gene
Affected Female
Unaffected Male
14
/
D/
D/
/
AUTOSOMAL DOMINANT
15
a/
a/
x/
/Y
x/
/Y
a/a
x/Y
RECESSIVE
RECESSIVE
X-LINKED
AUTOSOMAL
16
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17
Positional Cloning by Recombination Mapping
2. Follow which DNAs are co-inherited
(linked) Use DNA sequences that differ among
individuals within a family- Polymorphisms.
18
X
2
3
Parent
X
Gamete
X
Child
19
A1
B1
C1
X
2
3
Parent
B2
C2
A2
A2
B1
C1
X
Gamete
B1
C1
A2
X
Child
20
Recombination Mapping
Measures distance between 2 sites on a chromosome
according to frequency of recombination
Distance between 2 DNA markers or Distance
between a disease gene and a DNA marker
21
Male
Female
X
A1 B1
A1 B3
A2 B2
A1 B4
Meiosis
Gametes
Gametes
Offspring
22
Female
A1 B1
w y

A2 B2
45
A1 B1
w y
49
A2 B2

2
A1 B2
w
Easy for Drosophila
y
A2 B1
4
Gametes
A1 / A1 white eyes B1 / B1 yellow body
23
No fixed proportional
Conversion between
Genetic distance (cM)
and
Physical distance (kb, Mb)
24
VNTR / STRP DETECTION
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RECOMBINANT
Heterozygosity for A and B
Recognize haplotype
Linkage (cis)
MEIOSIS
27
INFORMATIVE MEIOSIS
Ideally- unambiguous inheritance of mutation
and markers (requires heterozygosity for each
in parent) knowledge of which alleles linked in
parent (phase)
28
Meiosis
Meiosis
Meiosis
A2 B2
A1 B1
29
FAMILY A
30
Assign numbers to results of linkage
analysis to deal with non-ideal meioses to
sum data from many meioses in a family to sum
data from several families
31
If unlinked-
?
If linked and RF
1/2
?
Likelihood of R
1 -
?
Likelihood of NR
1/2
Family A has 1 recombinant and 5 Non-Recombinants
?
Likelihood, given linkage of
Or given unlinked-
5
?
. (1- )
?
?
L ( )
6
L (1/2) (1/2)
32
Z 3
Lod
q
33
FAMILY B
A1

A2
D
NR
NR
NR
NR
NR
R
R
R
R
R
R
NR
34
Family B- Disease gene may be linked to A1 or A2
Consider equally likely
50 chance Family B has 1 R and 5 NR
50 chance Family B has 5 R and 1 NR
35
Phase unknown
?
0.1 0.2 0.3 0.4 0.5
Z 0.28 0.32 0.22 0..08 0
36
Phase known
Phase unknown
?
0.1 0.2 0.3 0.4 0.5
Z 0.28 0.32 0.22 0..08 0
37
For family A with meioses 1, 2, 3, 4 ..
Z Z1 Z2 Z3 Z4 ..
For multiple families, A, B, C, D..
Z Z(A) Z(B) Z(C) Z(D) .
Assumption same gene responsible for disease
in all families Problem locus heterogeneity
38
Z 3
Lod
q
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40
LINKAGE DISEQUILIBRIUM
Many generations
41
ZOO BLOT
42
PCR test DNA segments
43
SSCP
44
Normal
D
F506
45
Testing for specific mutations
46
ARMS 3 mis-match of primer
47
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48
TaqMan
49
OLA
50
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51
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