Download Funkcionális genomika

Real-Time PCR I.
Pair the bands (2-6 on the gel) with the melting curves!
Real-Time PCR I.
5
2
6
3
4
Pair the bands (2-6 on the gel) with the melting curves!
Real-Time PCR II.
5
2
6
3
4
What can be said about the genotype of the 5 patient if the examined samples are STR
sequences ?
Real-Time PCR II.
5
2
6
3
4
2 types of STR in each pearson.
2nd, 6th: homozygote, many repeat
3rds, 4th: heterozygote
5th: homozygote, few repeat
Real-Time PCR III.
detection of point mutation
3 patients; „A” gene expression analysis
What is the conclusion?
Real-Time PCR gyakorlati alkalmazások I.
pontmutáció detektálása
3 patients; „A” gene expression analysis
What is the conclusion?
3 genotypes
Real-Time PCR III.
detection of point mutation
3 patients; „A” gene expression analysis
What is the conclusion?
3 genotypes
Homozygote
wild type
Real-Time PCR III.
detection of point mutation
Homozygote
wild type
3 patients; „A” gene expression analysis
Heterozygote
What is the conclusion?
3 genotypes
Real-Time PCR III.
detection of point mutation
Homozygote
wild type
3 patients; „A” gene expression analysis
Homozygote mutant
Heterozygote
What is the conclusion?
3 genotypes
Healthy
Diseased
What is the correlation between the disease and the rate of gene expression?
Healthy
Diseased
Decreased mRNA copy number in diseased patient.
Healthy
Diseased
Setting of the threshold cycle
Healthy
Diseased
Healthy
Diseased
Define Ct value!
Healthy
Diseased
Healthy
Disease
Healthy: 21
Diseased: 31
∆Ct=2 (Ctb-Cte)
DNA chip
measuring gene expression
What do you see?
A
B
C
D
v
0h 2h 4h 6h 8h 10h
DNA chip
measuring gene expression
A gene continous, steady state expression (house keeping gene), B gene decreases, C increases,
D no expression
A
B
C
D
v
0h 2h 4h 6h 8h 10h
Measuring similarity between
expression patterns
Gene
name
0h
2h
4h
6h
8h
12h
Q
1
2
3
4
3
2
S
1
3
4
4
3
2
T
1
2
3
4
3
2
V
4
6
8
6
4
2
Q and T genes similar because of the same ratios at each time point.
How similar it its response to that of genes S and Q?
Measuring similarity between
expression patterns
Gene
name
0h
2h
4h
6h
8h
12h
Q
1
2
3
4
3
2
S
1
3
4
4
3
2
T
1
2
3
4
3
2
V
4
6
8
6
4
2
Pearson-correlation coefficient
It quantifies the extent to which the expression patterns of
two genes go up together and down together over several
time points.
Measuring similarity between
expression patterns
Gene
name
0h
2h
4h
6h
8h
12h
Q
1
2
3
4
3
2
S
1
3
4
4
3
2
T
1
2
3
4
3
2
V
4
6
8
6
4
2
Pearson-correlation coefficient
It quantifies the extent to which the expression patterns of two genes go
up together and down together over several time points.
=1: expression patterns of the 2 genes track perfectly
=-1: expression patterns of the 2 genes track perfectly, but in opposition to
the another.
0: expression patterns of the two genes do not track each other at all.
Measuring similarity between
expression patterns
Gene
name
0h
2h
4h
6h
8h
12h
Q
1
2
3
4
3
2
S
1
3
4
4
3
2
T
1
2
3
4
3
2
V
4
6
8
6
4
2
Find the correlation between Q and S genes:
1: compute the sample mean and sample standard
deviation of the expression values of each genes.
Xs=2,83 SS=1,067 Xq= 2,5 Sq= 0,957
Measuring similarity between
expression patterns
Gene
name
0h
2h
4h
6h
8h
12h
Q
1
2
3
4
3
2
S
1
3
4
4
3
2
T
1
2
3
4
3
2
V
4
6
8
6
4
2
Find the correlation between Q and S genes:
2: substract Xs from each value in the S row and divide each result by Ss.
Do the same in the Q row, to produce the following normalized row.
Snorm: -1,7; 0,16; 1,1; 1,1; 0,16; -0,5
Qnorm: -1,5; -0,5; 0,5; 1,5; 0,5; -0,5
Measuring similarity between
expression patterns
Gene
name
0h
2h
4h
6h
8h
12h
Q
1
2
3
4
3
2
S
1
3
4
4
3
2
T
1
2
3
4
3
2
V
4
6
8
6
4
2
Find the correlation between Q and S genes:
Multiply the first number in Snorm by the first number in Q norm, the
second number in Snorm….. And so on, keeping a running sum of these
products. Divide this sum by the number of elements in each row (6) to
get the correlation coefficient.
Measuring similarity between
expression patterns
Gene
name
0h
2h
4h
6h
8h
12h
Q
1
2
3
4
3
2
S
1
3
4
4
3
2
T
1
2
3
4
3
2
V
4
6
8
6
4
2
Find the correlation between Q and S genes:
ρ(Q,S)=0,897
Measuring similarity between
expression patterns
Gene
name
0h
2h
4h
6h
8h
12h
Q
1
2
3
4
3
2
S
1
3
4
4
3
2
T
1
2
3
4
3
2
V
4
6
8
6
4
2
Home work: find the correlation numbe r between T and V genes
Ρ (T,V)= -1