‹#›
=
Institute of Biology and Medical Genetics=
1st
F .M. and VFN, Praha
Molecular genetics II
=
![](3D"slide0022_image002.gif")
![](3D"slide0022_image004.gif")
![](3D"slide0022_image005.gif")
winter
semester
4th
week (October 27th – 3=
1st,
2008)
![](3D"slide0041_image006.gif")
DNA diagnostics
prenatal=
presymptomatic<=
/span>
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0041_image007.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
Clinical diagnosis
confirmation
Carrier detection
![](3D"slide0041_image008.gif")
![](3D"slide0041_image009.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
According to the targeting in the genome:
![](3D"slide0041_image010.gif")
Direct
DNA diagnostics
Determination of the allele (mutation) =
responsible for the disease (syndrome)
<=
/span>
=
Indirect
DNA diagnostics
Determination
of an allele (marker) that shows genetic l=
inkage
to the disease locus
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0042_image011.gif")
Direct DNA diagnostics
=
We are
searching for an allele (variant, mutation) that is responsible for the=
disease
phenotype in the family
=
![](3D"slide0042_image012.gif")
A mutat=
ion in
the disease gene identified in a parent
![](3D"slide0042_image013.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
=
![](3D"slide0042_image014.gif")
We are searching for that mutation in =
the
fetus
For example, in AD =
disease,
prenatal diagnosis:
![](3D"slide0042_image015.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0043_image016.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
=
Indirect DNA diagnostics
=
It is
not necessary to know, which mutation in the gene is <=
span
lang=3DCS style=3D'font-size:250%;text-shadow:auto'>responsible for the
disease, given we know, where the gene is =
localized on a chro=
mosome
![](3D"slide0043_image018.gif")
=
We use
genetic linkage between known marker that is placed alo=
ng =
the chromosome in v=
icinity
to the disease gene (known or unknown) with unknown causative mutation<=
/b> in the gene.
=
<=
span
lang=3DCS style=3D'font-size:250%;text-shadow:auto'>Which marker allele
cosegregates with the disease? Has the person in question =
got this
allele?
mutation
marker<=
/b>
gene
chromosome
Marker alleles typed in pa=
rents
Several markers are=
usually
assayed
![](3D"slide0043_image019.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
=
![](3D"slide0043_image020.gif")
Marker
alleles typed in offspring
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0044_image022.gif")
MAIN PI=
TFALL of
indirect DNA diagnostics: In different individuals, different marker alleles a=
re
linked to the mutated disease allele.
Vice ve=
rsa, the
same marker allele may be once linked to the =
disease allele, once to the
standard allele.
Therefo=
re, the
marker may be not informativ=
e, or
may be only partially infomative
=
A minor
difficulty – a possible recombination between the marker =
and the gene. Howev=
er, the
probability is small and can be estimated with reasonable accuracy.
Indirect DNA diagnostics
=
It is
not necessary to know, which mutation in the gene is <=
span
lang=3DCS style=3D'font-size:250%;text-shadow:auto'>responsible for the
disease, given we know, where the gene is =
localized on a chro=
mosome
=
We use
genetic linkage between known marker that is placed alo=
ng =
the chromosome in v=
icinity
to the disease gene (known or unknown) with unknown causative mutation<=
/b> in the gene.
=
<=
span
lang=3DCS style=3D'font-size:250%;text-shadow:auto'>Which marker allele
cosegregates with the disease? Has the person in question =
got this
allele?
![](3D"slide0045_image024.gif")
Variability on DNA level=
=
In the
population, certain piece of DNA seauence exists in several forms (alle=
les)
=
The
variable site may be situated in a gene (in exons – coding sequence, in introns
– noncoding sequence) or outside genes (i .e. always nonco=
ding
sequence)
Sorting according the allele frequency:
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0045_image025.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0045_image026.gif")
polym=
orfism
The
population frequency of the less common allele i=
s &g=
t; 1%=
<=
/span>
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
̶=
2;mutation“,
„rare allele“
![](3D"slide0045_image027.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
Population
frequency of such allele is < <=
/span>1%
Sorting according the functional impact:
funct=
ional
variant
The
allele influences the phenotype (at least in some
combination (recessive alleles)
<=
/span>
=
silent
variant
The
allele has no functional consequence=
<=
/span>
=
![](3D"slide0046_image028.gif")
Variability on DNA level
Types (=
that can
be assayed):
SNPs
Single nucleotide polymorp=
hisms
=
tande=
m repeats
Mikrosatellites, also dubbed STRs =3D short =
=
tandem
repeats; minisatellites
Str=
uctural polymorfisms
Insertions, deletions, inversions (ranging from
1 bp to chromosome)
=
Dup=
lications,
„low
copy repeats“
Actually a kind of tandem repeats, but differing
by detection techniques and funct=
ional significance;
e.g. =
FCGR3B and glomerulonephritis =
risk, RhD a RhCE , red and green opsins on
chromosome X, etc.
=
SNPs – single
TAG=
CCATCGGTANGTACTCAATGATCAGCT
A
![](3D"slide0036_image029.gif")
=
T
G
![](3D"slide0036_image030.gif")
C
![](3D"slide0036_image034.gif")
SNP
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
C
/ A
![](3D"slide0036_image038.gif")
99.9% DNA sequence is identical between any two chromosomes
(individuals). Of the remaining 0.1% difference, more than 80% is represen=
ted
by SNPs. It is tech=
nically
feasible now to type 1 000 000 SNPs in a single DNA sample, which should
expedite identification of the alleles responsible for a wide range of com=
mon
diseases.
![](3D"slide0036_image039.gif")
=
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
=
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
A
„special“ subset of SNPs:
RFLP =
=3D
restriction fragment length polymorphism
Restri=
ction
endonucleases are bacterial enzymes that cleave DNA at a specific sequence=
. <=
span
lang=3DCS style=3D'font-size:200%'>They protect bacteria against bacteriop=
hage
infection (or perhaps have no function)
=
4
Methods for SNP detection
TAGCCATCGGTACGTACTCAATGATCA
TAGCCATCGGTAAGTACTCAATGATCA
ATCGGTAGCCATGCATGAGTTACTAGT
ATCGGTAGCCATTCATGAGTTACTAGT
![](3D"slide0039_image040.gif")
MaeII
![](3D"slide0039_image041.gif")
![](3D"slide0039_image042.gif")
![](3D"slide0024_image043.gif")
=
Eco RI
Nomenclature of the restriction endonucle=
ases
![](3D"slide0024_image044.gif")
Three-letter organism abbreviation
„E scherichia
coli“
<=
/b>
„
More detailed data on the origin
R =3D enzyme from strain RY13,determined
by R-plasmid, number I
<=
/b>
R =3D enzyme from strain RY13,
![](3D"slide0024_image046.gif")
http://rebase.n=
eb.com
5
![](3D"slide0002_image047.gif")
=
![](3D"slide0002_image048.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0002_image049.gif")
=
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
Taq I 5´T C G A3&acu=
te; …=
5´A C G G T C G
A A T T3´
Taq I
5´T C G A<=
span
lang=3DCS style=3D'font-size:233%;text-shadow:auto;position:relative;top:-=
.45em;
mso-text-raise:30%'>3´ … 5´A C G G T C G A A T T3´
3´A G C T5´ …=
3´T G C C A G C T T A=
A5´
Hae III
5´G G C C 3´
C
C G G
=
Bam HI &n=
bsp;
5´G <=
/span>| G A T C C3´
![](3D"slide0002_image050.gif")
![](3D"slide0002_image051.gif")
![](3D"slide0002_image052.gif")
![](3D"slide0002_image054.gif")
Each restriction endonuclease recognizes =
and R=
11;
restriction site
6
![3Decori](3D"slide0023_image055.gif")
Cleavage of dsDNA by restriction endonucl=
ease
EcoRI
7
![](3D"slide0003_image056.gif")
restriction
si=
te
Alu I<=
b>
AG / CT
Sau 3AI <=
/b>
/ GTAC
Msp I<=
b>
C / CGG
5´G.G.G.C.G.T.A.C.A.T.A.G.C.T.A.A.T.G.G.C.A.A.G.C.=
T.A.T.G.G.T.3=
´
Find the restriction=
sites for the given restr=
iction
endonucleases:
![](3D"slide0003_image057.gif")
=
![](3D"slide0003_image058.gif")
![](3D"slide0003_image059.gif")
![](3D"slide0003_image060.gif")
![](3D"slide0003_image061.gif")
![](3D"slide0003_image062.gif")
![](3D"slide0003_image063.gif")
![](3D"slide0003_image064.gif")
![](3D"slide0003_image065.gif")
![](3D"slide0003_image066.gif")
=
Identification of the restriction sites
8
![](3D"slide0025_image067.gif")
C / CGG
Msp I<=
b>
/ GTAC
Sau 3AI <=
/b>
AG / CT
Alu I<=
b>
=
restriction
site
5´G.G.G.C.G.T.A.C.A.T.A.G.C.T.A.A.T.G.G.C.A.A.G.C.=
T.A.T.G.G.T.3=
´
![](3D"slide0025_image071.gif")
2x
1x=
3;
0
9
Identification of the restriction sites
Find the restriction=
sites for the given restr=
iction
endonucleases:
![](3D"slide0005_image072.gif")
Which of the given enzymes ca=
n be
used:
a) for cutting the given circ=
ular
DNA to linearize it?
b) for cutting out a fragment
containing a poly-A sequence?
G / GATCC
Bam HI=
CGAT / CG
Pvu I<=
b>
T / CGA
Taq I<=
b>
=
restriction
site
![](3D"slide0005_image073.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0005_image074.gif")
![](3D"slide0005_image075.gif")
![](3D"slide0005_image076.gif")
![](3D"slide0005_image077.gif")
=
![](3D"slide0005_image078.gif")
![](3D"slide0005_image079.gif")
![](3D"slide0005_image080.gif")
![](3D"slide0005_image081.gif")
![](3D"slide0005_image082.gif")
![](3D"slide0005_image083.gif")
2 sites - excission
![](3D"slide0005_image084.gif")
10
Identification of the restriction sites
![](3D"slide0010_image086.gif")
![](3D"slide0010_image087.gif")
Southern blot ana=
lysis
Description of the m=
ethod
![](3D"slide0010_image088.gif")
=
http://www.ac-creteil.fr/biotechnologies/doc_southernblotting.htm
11
Southern blot – animation
external file
external file
![](3D"slide0009_image089.gif")
![](3D"slide0009_image090.gif")
![](3D"slide0009_image091.gif")
![](3D"slide0009_image092.gif")
![](3D"slide0009_image093.gif")
DNA
probe
heating
(denaturation)<=
/span>
cooling
(renaturation,
=
hybridization)<=
/span>
![](3D"slide0009_image094.gif")
![](3D"slide0009_image095.gif")
![](3D"slide0009_image096.gif")
![](3D"slide0009_image097.gif")
Southern blot – probe hybridization=
12
![](3D"slide0011_image098.gif")
Task 4, p. 114 <=
span
lang=3DCS style=3D'font-size:300%;text-shadow:auto;color:blue;mso-color-in=
dex:
0'>Gene G
![](3D"slide0011_image099.gif")
![](3D"slide0011_image100.gif")
![](3D"slide0011_image101.gif")
![](3D"slide0011_image102.gif")
![](3D"slide0011_image103.gif")
![](3D"slide0011_image104.gif")
![](3D"slide0011_image105.gif")
![](3D"slide0011_image106.gif")
![](3D"slide0011_image107.gif")
![](3D"slide0011_image108.gif")
![](3D"slide0011_image109.gif")
![](3D"slide0011_image110.gif")
![](3D"slide0011_image111.gif")
![](3D"slide0011_image112.gif")
![](3D"slide0011_image113.gif")
![](3D"slide0011_image114.gif")
![](3D"slide0011_image115.gif")
![](3D"slide0011_image116.gif")
![](3D"slide0011_image117.gif")
![](3D"slide0011_image118.gif")
![](3D"slide0011_image119.gif")
![](3D"slide0011_image120.gif")
![](3D"slide0011_image121.gif")
![](3D"slide0011_image122.gif")
![](3D"slide0011_image123.gif")
![](3D"slide0011_image124.gif")
![](3D"slide0011_image125.gif")
1 kb
5´
3´
![](3D"slide0011_image126.gif")
=
![](3D"slide0011_image127.gif")
![](3D"slide0011_image128.gif")
![](3D"slide0011_image129.gif")
![](3D"slide0011_image130.gif")
![](3D"slide0011_image131.gif")
Taq I
EcoR I=
*
![](3D"slide0011_image132.gif")
=
![](3D"slide0011_image133.gif")
probe:
![](3D"slide0011_image134.gif")
=
cDNA
genomic
=
13
![](3D"slide0032_image161.gif")
1 kb
5´
3´
![](3D"slide0032_image162.gif")
=
![](3D"slide0032_image163.gif")
![](3D"slide0032_image164.gif")
Taq I
*
=
Results:
a) 4 or 5 fragments: 3kb, 8kb, 6kb/2kb+4kb, 7kb
![](3D"slide0032_image165.gif")
5´
3´
![](3D"slide0032_image166.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0032_image167.gif")
![](3D"slide0032_image168.gif")
![](3D"slide0032_image169.gif")
![](3D"slide0032_image170.gif")
![](3D"slide0032_image171.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0032_image172.gif")
=
![](3D"slide0032_image181.gif")
14
Task 4, p. 114 <=
span
lang=3DCS style=3D'font-size:300%;text-shadow:auto;color:blue;mso-color-in=
dex:
0'>Gene G, cleavage
![](3D"slide0034_image182.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
=
=
=
![](3D"slide0034_image209.gif")
1 kb
5´
3´
![](3D"slide0034_image210.gif")
=
![](3D"slide0034_image211.gif")
![](3D"slide0034_image212.gif")
Taq I
*
=
Results:
![](3D"slide0034_image213.gif")
5´
3´
![](3D"slide0034_image214.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0034_image215.gif")
![](3D"slide0034_image216.gif")
![](3D"slide0034_image217.gif")
![](3D"slide0034_image218.gif")
![](3D"slide0034_image219.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0034_image220.gif")
=
b)&nb=
sp;
3
fragments: 3kb, 8kb, 7kb
c) no
15
Task 4, p. 114 <=
span
lang=3DCS style=3D'font-size:300%;text-shadow:auto;color:blue;mso-color-in=
dex:
0'>Gene G, Southern blot with a cDNA probe
![](3D"slide0035_image230.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0035_image257.gif")
1 kb
5´
3´
![](3D"slide0035_image258.gif")
=
![](3D"slide0035_image259.gif")
![](3D"slide0035_image260.gif")
Taq I
*
=
Results:
![](3D"slide0035_image261.gif")
5´
3´
![](3D"slide0035_image262.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0035_image263.gif")
![](3D"slide0035_image264.gif")
![](3D"slide0035_image265.gif")
![](3D"slide0035_image266.gif")
![](3D"slide0035_image267.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0035_image268.gif")
=
d) Yes 4kb (alle=
le
+) 6kb (allele -)<=
/span>
16
Task 4, p. 114 <=
span
lang=3DCS style=3D'font-size:300%;text-shadow:auto;color:blue;mso-color-in=
dex:
0'>Gene G, Southern blot with a genomic probe=
![](3D"slide0026_image278.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
=
![](3D"slide0026_image279.gif")
![](3D"slide0026_image280.gif")
![](3D"slide0026_image281.gif")
![](3D"slide0026_image282.gif")
![](3D"slide0026_image283.gif")
![](3D"slide0026_image284.gif")
![](3D"slide0026_image285.gif")
![](3D"slide0026_image286.gif")
![](3D"slide0026_image287.gif")
![](3D"slide0026_image288.gif")
![](3D"slide0026_image289.gif")
![](3D"slide0026_image290.gif")
![](3D"slide0026_image291.gif")
![](3D"slide0026_image292.gif")
![](3D"slide0026_image293.gif")
![](3D"slide0026_image294.gif")
=
M
![](3D"slide0026_image295.gif")
![](3D"slide0026_image296.gif")
![](3D"slide0026_image297.gif")
=
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0026_image298.gif")
![](3D"slide0026_image299.gif")
![](3D"slide0026_image300.gif")
![](3D"slide0026_image301.gif")
![](3D"slide0026_image302.gif")
![](3D"slide0026_image303.gif")
![](3D"slide0026_image304.gif")
![](3D"slide0026_image305.gif")
![](3D"slide0026_image306.gif")
![](3D"slide0026_image307.gif")
![](3D"slide0026_image308.gif")
![](3D"slide0026_image309.gif")
![](3D"slide0026_image310.gif")
![](3D"slide0026_image311.gif")
![](3D"slide0026_image312.gif")
=
P
![](3D"slide0026_image313.gif")
E1 – restriction endonu=
clease
1
E2 – restriction endonu=
clease
2
M – fragment of maternal
chromosome
P – fragment of paternal
chromosome
¯ - restriction site
(restriction endonuclease cleaves DNA here)
2cM – linkage map dista=
nce
between gene and extragene probe
![](3D"slide0026_image314.gif")
=
![](3D"slide0026_image315.gif")
=
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
Extragenic probe
17
![](3D"slide0030_image316.gif")
=
M
=
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0030_image317.gif")
E1
![](3D"slide0030_image318.gif")
E1
![](3D"slide0030_image319.gif")
E1
![](3D"slide0030_image320.gif")
E1
![](3D"slide0030_image321.gif")
E1
![](3D"slide0030_image322.gif")
E1
![](3D"slide0030_image323.gif")
E1
![](3D"slide0030_image324.gif")
![](3D"slide0030_image325.gif")
![](3D"slide0030_image326.gif")
=
P
![](3D"slide0030_image327.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
Extragenic probe – cutting with E1
enzyme
Although restriction
endonuclease E1 will cut DNA at all restriction sites, we will be able to detect on the autora=
diogram
only fragments labeled by the hybridized probe. These are of the same lengt=
h on
both chromosomes, the
individual is thus a homozygote in restriction fragment length.
![](3D"slide0030_image328.gif")
=
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0030_image329.gif")
E1
![](3D"slide0030_image330.gif")
E1
![](3D"slide0030_image331.gif")
E1
![](3D"slide0030_image332.gif")
E1
![](3D"slide0030_image333.gif")
E1
![](3D"slide0030_image334.gif")
E1
![](3D"slide0030_image335.gif")
E1
![](3D"slide0030_image336.gif")
![](3D"slide0030_image337.gif")
![](3D"slide0030_image338.gif")
![](3D"slide0030_image339.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0030_image340.gif")
![](3D"slide0030_image341.gif")
E1
=
autoradiogram=
18
![](3D"slide0031_image342.gif")
=
M
=
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0031_image343.gif")
E2
![](3D"slide0031_image344.gif")
E2
![](3D"slide0031_image345.gif")
E2
![](3D"slide0031_image346.gif")
E2
![](3D"slide0031_image347.gif")
E2
![](3D"slide0031_image348.gif")
![](3D"slide0031_image349.gif")
![](3D"slide0031_image350.gif")
=
P
![](3D"slide0031_image351.gif")
E2
![](3D"slide0031_image352.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0031_image353.gif")
=
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0031_image354.gif")
E2
![](3D"slide0031_image355.gif")
E2
![](3D"slide0031_image356.gif")
E2
![](3D"slide0031_image357.gif")
E2
![](3D"slide0031_image358.gif")
E2
![](3D"slide0031_image359.gif")
![](3D"slide0031_image360.gif")
![](3D"slide0031_image361.gif")
![](3D"slide0031_image362.gif")
E2
![](3D"slide0031_image363.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0031_image364.gif")
![](3D"slide0031_image365.gif")
![](3D"slide0031_image366.gif")
E1
E2
=
autoradiogram=
E2
=
19
Extragenic probe – cutting with E2
enzyme
Although the restric=
tase E2
will cut DNA at all restriction sites, we will be able to detect only the fragments label=
ed by
the hybridized probe. The fragments are of different length on maternal and
paternal chromosomes, the
individual is thus a
heterozygote for the length of restriction fragments.
![](3D"slide0014_image367.gif")
![](3D"slide0014_image368.gif")
![](3D"slide0014_image369.gif")
=
![](3D"slide0014_image370.gif")
=
![](3D"slide0014_image371.gif")
=
![](3D"slide0014_image372.gif")
![](3D"slide0014_image373.gif")
![](3D"slide0014_image374.gif")
![](3D"slide0014_image375.gif")
![](3D"slide0014_image376.gif")
![](3D"slide0014_image377.gif")
![](3D"slide0014_image378.gif")
![](3D"slide0014_image379.gif")
![](3D"slide0014_image380.gif")
![](3D"slide0014_image381.gif")
=
1
The
family is noninformative. All family members are homozygotes in length of the
restriction fragments.=
(the parents are heterozygotes in the gene of interest). However, we do not
know, which allele of the gene was transmitted with allele 1 of the RFLP.<=
/b>
![](3D"slide0014_image382.gif")
=
![](3D"slide0014_image383.gif")
![](3D"slide0014_image384.gif")
=
![](3D"slide0014_image385.gif")
![](3D"slide0014_image386.gif")
=
![](3D"slide0014_image387.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0014_image388.gif")
![](3D"slide0014_image389.gif")
![](3D"slide0014_image390.gif")
![](3D"slide0014_image391.gif")
![](3D"slide0014_image392.gif")
=
1
![](3D"slide0014_image393.gif")
=
2
This
family is only partially informative. It is not possible to disting=
uish
the origin of RFLP fragment with mutant allele between mother and father.<=
/b> However, the second child is a homozygote (AA or=
aa)
20
![](3D"slide0015_image394.gif")
![](3D"slide0015_image395.gif")
![](3D"slide0015_image396.gif")
=
![](3D"slide0015_image397.gif")
![](3D"slide0015_image398.gif")
=
![](3D"slide0015_image399.gif")
![](3D"slide0015_image400.gif")
![](3D"slide0015_image401.gif")
![](3D"slide0015_image402.gif")
![](3D"slide0015_image403.gif")
![](3D"slide0015_image404.gif")
=
1
![](3D"slide0015_image405.gif")
=
2
=
![](3D"slide0015_image406.gif")
=
=
21
![](3D"slide0016_image407.gif")
![](3D"slide0016_image408.gif")
![](3D"slide0016_image409.gif")
![](3D"slide0016_image410.gif")
=
![](3D"slide0016_image411.gif")
![](3D"slide0016_image412.gif")
![](3D"slide0016_image413.gif")
![](3D"slide0016_image414.gif")
![](3D"slide0016_image415.gif")
![](3D"slide0016_image416.gif")
![](3D"slide0016_image417.gif")
=
1
![](3D"slide0016_image418.gif")
![](3D"slide0016_image419.gif")
=
2
![](3D"slide0016_image420.gif")
=
![](3D"slide0016_image421.gif")
![](3D"slide0016_image422.gif")
![](3D"slide0016_image423.gif")
![](3D"slide0016_image424.gif")
![](3D"slide0016_image425.gif")
![](3D"slide0016_image426.gif")
![](3D"slide0016_image427.gif")
=
![](3D"slide0016_image428.gif")
=
![](3D"slide0016_image429.gif")
![](3D"slide0016_image430.gif")
![](3D"slide0016_image431.gif")
![](3D"slide0016_image432.gif")
![](3D"slide0016_image433.gif")
![](3D"slide0016_image434.gif")
=
1
=
2
<=
b>The
affected son (1,2) has inherited fragment 1 from his father (1,1) and frag=
ment
2 from mother (1,2) fragment 2, which is linked to the mutant allele. The
second child (1,1) has maternal fragment 1 with the normal allele and frag=
ment
1 from father. We don´t know if this fragment carries mutant or norm=
al
allele. So the prognosis is 50% carrier, 50% healthy homozygote=
![](3D"slide0016_image435.gif")
=
![](3D"slide0016_image436.gif")
![](3D"slide0016_image437.gif")
![](3D"slide0016_image438.gif")
22
![](3D"slide0017_image439.gif")
Are children II/2 and II/3
heterozygotes for the mutant allele?
AR disease, intragene probe (complete linkage)
![](3D"slide0017_image440.gif")
![](3D"slide0017_image441.gif")
![](3D"slide0017_image442.gif")
![](3D"slide0017_image443.gif")
![](3D"slide0017_image444.gif")
![](3D"slide0017_image445.gif")
![](3D"slide0017_image446.gif")
1
![](3D"slide0017_image447.gif")
![](3D"slide0017_image448.gif")
![](3D"slide0017_image449.gif")
2
![](3D"slide0017_image450.gif")
![](3D"slide0017_image451.gif")
![](3D"slide0017_image452.gif")
I
II
![](3D"slide0017_image453.gif")
=
1
![](3D"slide0017_image454.gif")
=
1
![](3D"slide0017_image455.gif")
=
2
![](3D"slide0017_image456.gif")
=
2
3
aa
AA
![](3D"slide0017_image457.gif")
a
![](3D"slide0017_image458.gif")
a
![](3D"slide0017_image459.gif")
A
![](3D"slide0017_image460.gif")
A
Task 1, p. 120<=
/span>
23
![](3D"slide0018_image461.gif")
![](3D"slide0018_image462.gif")
![](3D"slide0018_image463.gif")
![](3D"slide0018_image464.gif")
![](3D"slide0018_image465.gif")
![](3D"slide0018_image466.gif")
![](3D"slide0018_image467.gif")
![](3D"slide0018_image468.gif")
1
![](3D"slide0018_image469.gif")
![](3D"slide0018_image470.gif")
![](3D"slide0018_image471.gif")
2
![](3D"slide0018_image472.gif")
![](3D"slide0018_image473.gif")
![](3D"slide0018_image474.gif")
I
II
![](3D"slide0018_image475.gif")
=
1
![](3D"slide0018_image476.gif")
=
1
![](3D"slide0018_image477.gif")
=
2
![](3D"slide0018_image478.gif")
=
2
3
aa
AA
![](3D"slide0018_image479.gif")
a
=
![](3D"slide0018_image480.gif")
a
![](3D"slide0018_image481.gif")
A
=
![](3D"slide0018_image482.gif")
A
II/2 no
II/3 yes
24
Task 1, p. 120<=
/span>
Are children II/2 and II/3
heterozygotes for the mutant allele?
AR disease, intragene probe (complete linkage)
![](3D"slide0019_image483.gif")
Task 2, p. 120<=
/span>
![](3D"slide0019_image484.gif")
![](3D"slide0019_image485.gif")
![](3D"slide0019_image486.gif")
![](3D"slide0019_image487.gif")
![](3D"slide0019_image488.gif")
=
1
![](3D"slide0019_image489.gif")
![](3D"slide0019_image490.gif")
=
2
![](3D"slide0019_image491.gif")
![](3D"slide0019_image492.gif")
![](3D"slide0019_image493.gif")
=
I
=
II
![](3D"slide0019_image494.gif")
=
![](3D"slide0019_image495.gif")
=
![](3D"slide0019_image496.gif")
=
![](3D"slide0019_image497.gif")
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
![](3D"slide0019_image498.gif")
II/2 AA=
or Aa
II/3 AA=
or Aa=
=
Heterozygosity of I=
I/2 and
II/3 is undeterminable - RFLP analysis =
is unsuccessful=
.
![](3D"slide0019_image499.gif")
Aa
Aa
a
a
25
Are children II/2 and II/3
heterozygotes for the mutant allele?
AR disease, intragene probe (complete linkage)
![](3D"slide0020_image500.gif")
<=
/div>
![](3D"slide0020_image501.gif")
![](3D"slide0020_image502.gif")
![](3D"slide0020_image503.gif")
![](3D"slide0020_image504.gif")
![](3D"slide0020_image505.gif")
![](3D"slide0020_image506.gif")
![](3D"slide0020_image507.gif")
=
1
![](3D"slide0020_image508.gif")
=
2
![](3D"slide0020_image509.gif")
![](3D"slide0020_image510.gif")
![](3D"slide0020_image511.gif")
=
I
=
II
![](3D"slide0020_image512.gif")
=
![](3D"slide0020_image513.gif")
2
![](3D"slide0020_image514.gif")
Is daughter II/3 heterozygote for the mutant all=
ele
of haemophilia?
X-linked recessive disease, intragene probe
(complete linkage)
![](3D"slide0020_image515.gif")
X+Y
X+Xh=
XhY
X+Y
![](3D"slide0020_image516.gif")
Xh
![](3D"slide0020_image517.gif")
X+
X+
![](3D"slide0020_image518.gif")
X+
II/3 yes
![](3D"slide0020_image519.gif")
Xh
Xh
26
Task 3, p. 121<=
/span>
![](3D"slide0021_image520.gif")
![](3D"slide0021_image521.gif")
![](3D"slide0021_image522.gif")
![](3D"slide0021_image523.gif")
![](3D"slide0021_image524.gif")
![](3D"slide0021_image525.gif")
=
1
![](3D"slide0021_image526.gif")
=
2
![](3D"slide0021_image527.gif")
![](3D"slide0021_image528.gif")
![](3D"slide0021_image529.gif")
=
I
=
II
![](3D"slide0021_image530.gif")
=
1
![](3D"slide0021_image531.gif")
=
2
![](3D"slide0021_image532.gif")
3
=
2
<=
span
lang=3DCS style=3D'mso-special-format:lastCR;display:none'>
1
II/3 no
![](3D"slide0021_image533.gif")
Xh
Xh
X+
X+
X+
X+
X+
27
Task 4, p. 121<=
/span>
Is daughter II/3 heterozygote for the mutant all=
ele
of haemophilia?
X-linked recessive disease,
intragene probe <=
/div>
Klepnutím lze upravit styly předlohy textu.
=
Druhá úroveň
Třetí úroveň
Čtvrtá úroveň
Pátá úroveň
‹#›
10<=
/div>
11<=
/div>
' + ENDSHOW_MESG + '
![](3D"notes_flag.gif")
Po=
známky![](3D"buttons.gif")
![](3D"slide0022_image001.png"){kind=link}
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