welcome to introduction to bioinformatics monday, 21 march 2005 genome comparison coming attractions...

Welcome toIntroduction to Bioinformatics

Monday, 21 March 2005

Genome Comparison

• Coming attractions

• How to compare genomes

• Chi-squared analysis

E. coli: What makes it kill?

Escherichia coli . . .

. . . very small lab rats

Courtesy of Kent State University Microbiology


Escherichia coli . . .

haemorrhagic colitis


E. coli K12 E. coli O157:H7

TCTACTTATA TTCAATCCAC AGGGCTACACAAGAGTCTGT TGAATGAACA CATACATGGTTTCTGTCTGC TCTGACCTCT GGCAGCTTTC TGGATTTCGG AACTCTAGCC TGCCCCACTC GAACCTTAGT GACTTCTGCT ATACCAAAGT CTCCGTAAAC CTCTAACATG ATGTCAGCAA TGAATAAACT TTGTTAAAGG TACAAATGAA AAGAGTTTAA AGTTAAAAAC GAATTGCAGT AAACCTGTAT GGTTACATGA ACTGCCTAAA TTATATATTT TAAGAAATTA ATTGCAATTA CCCCAGCTGT CATTAAAAAG AGGCAAATAC GACAGCACTG ACCCTCAAGA AGGCACCGGC GCTGAAATTC CGCTGAGAGC AGAGTGGTAC CCCTGCACCA GGTCTTTCCT GTGGGCACTG ATGAATGACT GAACGAACGA TTGAATGAAA

TCTACTTATA TTCAATCCAC AGGGCTACACAAGAGTCTGT TGAATGAACA CATACATGGTTTCTGTCTGC TCTGACCTCT GGCAGCTTTC TGGATTTCGG AACTCTAGCC TGCCCCACTC GAACCTTAGT GACTTCTGCT ATACCAAAGT CTCCGTAAAC CTCTAACATG ATGTCAGCAA TGAATAAACT TTGTTAAAGG TACAAATGAA AAGAGTTTAA AGTTAAAAAC GAATTGCAGT AAACCTGTAT GGTTACATGA ACTGCCTAAA TTATATATTT TAAGAAATTA ATTGCAATTA CCCCAGCTGT CATTAAAAAG AGGCAAATAC GACAGCACTG ACCCTCAAGA AGGCACCGGC GCTGAAATTC CGCTGAGAGC AGAGTGGTAC CCCTGCACCA GGTCTTTCCT GTGGGCACTG ATGAATGACT GAACGAACGA TTGAATGAAA

How to compare genomes

E. coli O157:H7 genomeGATAGATCCCCACGCCTATAATGGCGCATAACACACTAAACTTGGGGTATTGAAGCAGTCGCCAAAGAGTGACCGGTCATCCTTCTCCGCTGCGAAATATCCTTCTTGTTGGCATACCACGCCTTGA...

E. coli K12 genomeGCGGAGCAAACTGGGCGTCTTTCGAGAACTAACAAATCCGATTGCGGGCTTCTCACGCATAGGCGCAGTTATGGTTAATGCCAAAACTTTTTTTTCGCGCCGAAATAACATAATGCACAGGCATGGC...



E. coli O157:H7 genome

GATAGATCCCCACGCCTATAATGGCGCATAACACACTAAACTTGGGGTATTGAAGCAGTCGCCAAACCACGCCTTGA...

GATAGATCCCC





CCCACGCCTAT

E

. col

i O15

7:H

7

E. coli K12

E

. col

i O15

7:H

7

E. coli K12

O-Islands

P

roch

lor

ss12

0

Prochlor. MED4

Prochlorococcus SS120

Prochlorococcus MED4

(100 nuc)

P

roch

lor

ss12

0

Prochlor. MED4

Prochlorococcus SS120

Prochlorococcus MED4

(25 nuc)

Nature of Pathogenicity Islands

Horizontal transfer of foreign DNA



How do differences arise between genomes?

InfectionPhage

Bacterial chromosome

Phage genome

Lysogenicpathway

LyticpathwayPhage genome

DeathGeneral transduction


InfectionPhage


Phage genome

Lysogenicpathway


Life!


InfectionPhage


Phage genome

Lysogenicpathway


Special transduction




• Differences in genome sequence Useful only if very related





• Differences in protein content Useful for even distant comparisons






How to find orthologous protein?






How to find corresponding protein?


X X X X

X

X

X

Yeast E. coli Anabaena Methanobacter


X X X X

X

X

X

Yeast E. coli Anabaena Methanobacter

All similar protein?

Most related by common descent?

Orthologs

OrthologsParalogs



All similar protein?OrthologsParalogsBlast

E-valuethreshold

Organism X

Organism Y



Orthologs

Blast

E-valuethreshold

Organism Y

Organism X

Organism Y

Organism Y

Defined by bidirectional Blast hit


PROTEINS-SIMILAR-TO

ORTHOLOG-OF

COMMON-ORTHOLOGS-OF

Nature of Pathogenicity IslandsNucleotide frequencies comparisons

Base Sequence1 Sequence2 Total

A 1,000 600 1,600

C 1,000 800 1,800

G 1,000 700 1,700

T 1,000 900 1,900

Total 4,000 3,000 7,000

Nucleotide Count

Nucleotide frequencies to detect foreign genes

1. Find nucleotide frequencies of native genes

2. Find nucleotide frequencies of test gene

3. Compare frequencies

4. How likely differences arose by chance?

Chi-squared analysis

Result: 705 purple 224 white = 929 plantsResult: 698 purple 231 white = 929 plantsResult: 688 purple 241 white = 929 plantsResult: 710 purple 219 white = 929 plantsResult: 695 purple 234 white = 929 plantsResult: 702 purple 227 white = 929 plants

Where does 2 come from?A million repetitions of Mendel’s experiment

Create a million universes -- purple:white on average = 3:1

200,000 repetitions


500,000 repetitions



1,000,000 repetitions

Why is it that the two dotted lines are on opposite sides of the mean?


1,000,000 repetitions

What’s the most likely result? How often does it occur?

Deviation from ExpectationTwo example experiments

Why is there shading on both sides of the curve?

The farther away O from E, the smaller/larger the shaded area?

Steps in Performing a Chi2 Test

Determine the expected values for the experiment

Model: 3 purple : 1 white flowerTotal counted: 929Purple = 75% of 929 = 696.75White = 25% of 929 = 232.25

Calculate the squares of the deviations

Chi2 = Sum of (O - E)2 / EChi2 = (705 - 696.75)2 /696.75 + (224 - 232.25)2 /232.25 ~82 / 700 + ~82 / 230 ~0.09 ~0.3 Chi2 = approx 0.39 (actually = 0.37)


Determine the degrees of freedom

What was the experiment? - Count 929 flowers a million timesAsk: purple? (if not, then white)

Look up probability for 2 value

2 = 0.30

80% > P > 50%. Call it ~60%

Therefore ONE degree of freedom


P ~60%

Draw a conclusion

The result has a 50% chance of being correctThe hypothesis has a 50% chance of being correct60% of the time, Mendel’s result or worse would have arisen by chance if purple:white truly occurs in a 3:1ratio.

Deviation from ExpectationTwo example experiments

Study Question 20:What if Mendel had counted not 929 but 929,000 plants -- what does the curve and shading look like then? (d still = 29)

P = .50 P = ???

Interpretation of Chi-Square

Does a high P value indicate the hypothesis is correct?

Does a low P value indicate the hypothesis is incorrect?

Bag of Marbles

1000’s of marbles!50% red, 50% blue

Guaranteed!

Test Claim of 50%:50%

41 marbles

59 marbles

100 marblesTOTAL

Is their claim correct?

How to tell how close is close enough?

2 Test of Claim

Chi2 = Sum of (O - E)2 / E

Chi2 = (53 - 50)2 /50 + (47 - 50)2 /50 9 / 50 + 9 / 50 18/50 0.36

P = ?P = ~60%

welcome to introduction to bioinformatics monday, 21 march 2005 genome comparison coming attractions...

Documents

escherichia coli

coli k12e

genomes chisquared analysise

genome comparison

haemorrhagic colitise