Summary of 8th lesson

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Summary of 8th lesson

• Exotic microbes have a reduced level of genetic
  variability
• If genotypes fall on clades separated by long
  branches, it may be an indication there is no sex
  going on between individuals belonging to the two
  branches. Formal tests like the Index of
  association can test for that
• Anonymous multilocus analysis can be done without
  any knowledge of the genome using markers such as
  RAPDs or AFLPs
• Need to eliminate co-segregant markers and to use
  Jaccards

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Dealing with dominant anonymous multilocus markers

• Need to use large numbers (linkage)
• Repeatability
• Graph distribution of distances
• Calculate distance using Jaccards similarity
  index

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Jaccards

• Only 1-1 and 1-0 count, 0-0 do not count
• 1010011
• 1001011
• 1001000

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Jaccards

• Only 1-1 and 1-0 count, 0-0 do not count
• A 1010011 AB 0.6 0.4 (1-AB)
• B 1001011 BC0.5 0.5
• C 1001000 AC0.2 0.8

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Now that we have distances.

• Plot their distribution (clonal vs. sexual)

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Now that we have distances.

• Plot their distribution (clonal vs. sexual)
• Analysis
• Similarity (cluster analysis) a variety of
  algorithms. Most common are NJ and UPGMA

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Now that we have distances.

• Plot their distribution (clonal vs. sexual)
• Analysis
• Similarity (cluster analysis) a variety of
  algorithms. Most common are NJ and UPGMA
• AMOVA requires a priori grouping

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AMOVA groupings

• Individuals within population
• Among populations
• Among regions
• AMOVA partitions molecular variance amongst a
  priori defined groupings

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Results Jaccard similarity coefficients
P. nemorosa
P. pseudosyringae U.S. and E.U.
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P. pseudosyringae genetic similarity patterns are
different in U.S. and E.U.
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Results P. nemorosa
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Have we sampled enough?

• Resampling approaches
• Saturation curves
• A total of 30 polymorphic alleles
• Our sample is either 10 or 20
• Calculate whether each new sample is
  characterized by new alleles

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Saturation curves
No Of New alleles
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20
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If we have codominant markers how many do I need

• IDENTITY tests probability calculation based
  on allele frequency Multiplication of
  frequencies of alleles
• 10 alleles at locus 1 P10.1
• 5 alleles at locus 2 P20,2
• Total P P1P20.02

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White mangroves Corioloposis caperata
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Distances between study sites
White mangroves Corioloposis caperata
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Forest fragmentation can lead to loss of gene
flow among previously contiguous populations.
The negative repercussions of such genetic
isolation should most severely affect highly
specialized organisms such as some
plant-parasitic fungi.
AFLP study on single spores
Coriolopsis caperata on Laguncularia racemosa
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Using DNA sequences

• Obtain sequence
• Align sequences, number of parsimony informative
  sites
• Gap handling
• Picking sequences (order)
• Analyze sequences (similarity/parsimony/exhaustive
  /bayesian
• Analyze output CI, HI Bootstrap/decay indices

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Using DNA sequences

• Testing alternative trees kashino hasegawa
• Molecular clock
• Outgroup
• Spatial correlation (Mantel)
• Networks and coalescence approaches

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Good chromatogram!
Bad chromatogram
Reverse reaction suffers same problems in
opposite direction
Pull-up (too much signal)
Loss of fidelity leads to slips, skips and mixed
signals
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Alignments (Se-Al)
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Distance vs. parsimony

• Distance simply calculates at how many positions
  sequences are similar or differen
• (Matteo) ACGTAACGTT-AG
• (Amanda) AGTTAACGTTAAG
• (Patrick) ACTTAACGTTAAG

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Distance vs. Parsimony
Amanda

• Patrick

Patrick
Amanda Matteo

• Matteo

OUTGROUP can allow us to pick Matteo as ancestral
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Confidence

• Bootstrap (resampling approcah) Decay indices
  (threshold approach)
• Consistency index
• Homoplasy index

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Pacifico
Caribe
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The scale of disease

• Dispersal gradients dependent on propagule size,
  resilience, ability to dessicate, NOTE not
  linear
• Important interaction with environment, habitat,
  and niche availability. Examples Heterobasidion
  in Western Alps, Matsutake mushrooms that offer
  example of habitat tracking
• Scale of dispersal (implicitely correlated to
  metapopulation structure)---

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From Garbelotto and Chapela, Evolution and
biogeography of matsutakes
Biodiversity within species as significant as
between species
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Other important types of markers (co-dominant)

• Restriction Fragment Length Polymorphisms (RFLP)
  of a locus
• Single Nucleotide Polymorphisms (SNPs)
• Microsatellites (SSR)

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Restriction Fragment Length Polymorphisms (RFLP)
of a locus

• aacccacgtcaataaaaa
• aacccac
• gtcaataaaaa
• One restriction site

aacccaggtcaataaaaa aacccaggtcaataaaaa No
restriction sites
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Restriction Fragment Length Polymorphisms (RFLP)
of a locus
Two alternate alleles codominant marker
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Single Nucleotide Polymorphisms (SNPs)

• nnACGTnnnnnnTAAGnnnnnn
• nnAGGTnnnnnnTATGnnnnnn

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Dept. of Energy / Joint Genome Institute (www.JGI.
gov)
Shotgun sequencing Completed May 2004 - 7x
coverage 66 MB..much larger than first
calculated Used 445,030 reads (FASTA format) -
3x coverage
FASTA format
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1. SSR detection
batch search di and tri-nucleotide
repeats differ in repeat length
CCGAAATCGGACCTTGAGTGCGGAGAGAGAGAGAGACTGTACGAGCCCGA
GTCTCGCAT
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locus ct 0070
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Microsatellites (SSR)

• Supposed to be neutral
• Stepwise mutation model
• Very sensitive because loci are prone to mutation
• Allele is af fragment of DNA that includes the
  flanking regions of the microsatellite and then a
  certain number of tandem repeats (variation in
  size should be in multiple of SSR(