What are microsatellites

1
What are microsatellites?

• Tandem repeats of 1-6 nucleotides that are 5 to
  40 repeats long
• Dinucleotide, trinucleotide, tetranucleotide
  repeats most common
• Found at high frequency in nuclear genomes of
  most taxa

2
Characteristics of microsatellite mutation

• Many have high mutation rate
• Rate 0.01 to 0.000001 mutations per locus per
  generation, average 0.0005
• Mutations occur due to slippage and proofreading
  errors during replications
• Mutations result in changes in length of repeat
  string
• Length differences can be seen w/ high
  resolution gel electrophoresis, which allows
  rapid genotyping
• High mutation rate generates allelic diverstiy
  necessary for genetic studies of processes acting
  on ecological time scales

3
Some questions addressed using microsatellite
variation

• Number of populations and which an individual
  originated from
• Data requirements
• Multilocus, allele frequency
• gt10 microsatelites, lower migration and smaller
  pop. size fewer loci
• Recent historical population expansion or
  contraction and size
• Data requirements
• Highly polymorphic microsatellite or sequence
  data
• gt 1 locus will dampen interlocus sampling error

4
Some questions addressed using microsatellite
variation

• Individuals genetic relationships, movement, and
  which are clones
• Data requirements
• Multilocus, genotype identification
• possible w/ microsatelites, AFLP and RAPD
• Dispersal dist., source sink relationships,
  extinction and recolonization, LS effect on
  structure and migration, ?s in structure and
  connectivity
• Data requirements
• Many marker types
• gt 1 locus will dampen interlocus sampling error

5
Pattern of allele diversity of cattle in
AfricaDeveloped using 183 microsatellite allele
frequencies
6
Pros of using microsatellites

• Sample preparation
  
• Small sample of DNA necessary (thank PCR)
• Relatively short length makes DNA degradation
  less problematic
• Probably not to same extent of SNPs?
• Only need cheap extraction kit
• Species specific primers reduce problem of
  cross contamination
• Contamination a problem in fecal samples and
  corals (symbiotic algae)
• Does this apply during primer preparation?

7
Pros of using microsatellites

• Information content
• Multilocus technique
• Reduces sampling error
• Greater statistical power
• Codominant markers
• Can identify heterozygote
• Less expensive per loci
• Compared to sequencing multiple loci
• Multiple loci can be genotyped simultaneously
• High mutation rate
• Allows questions of finer timescales to be
  addressed (10-100 generations)
• High loci diversity important when addressing
  certain questions

8
Cons of using microsatellites

• Complex mutational processes
• Creates problems when using Fst and Rst
  statistics
• Metrics employing stepwise mutation model are
  sensitive to violation of SSM
• Hidden allelic diversity complicates size based
  allele identification
• Homoplasy, detectable by sequencing for base
  pair substitution, but not if by back-mutation
• Amplification problems
• High mutation rate in flanking region lead to
  problems with markers
• Null alleles
• Repeat regions rare in invertebrates,
  lepidopterans, and birds

9
Acquiring microsatellite markers

• Literature
• Molecular Ecology Notes
• Searchable databases
• http//tomato.bio.trinity.edu/
• GenBank searchable through National center of
  Biotech. Info.
• Cross-species amplification
• Possible when flanking regions highly conserved

10
New markers

• Many labs offer isolation services
• Should conduct screening to optimize set of loci
• Violating assumptions could lead to inaccurate
  and biased estimates
• Assumptions
• Accurately scored genotypes
• All alleles amplified
• Linkage equilibrium
• Hardy-Weinburg equilibrium
• Selective neutrality
• Mendelian inheritance
• All alleles differ in length

11
Common repeat sequences
12
Traditional vs contemporary methods
13
References

• Manel, S., M. K. Schwartz, G. Luikart, and P.
  Taberlet. 2003. Landscape genetics combining
  landscape ecology and population genetics. TREE
  18(4) 189-197.
• Selkoe, K. A., and R. J. Toonen. 2006.
  Microsatellites for ecologists a practical guide
  to using and evaluating microsatellite markers.
  Ecology Letters 9 615-629.
• Zane, L., L., Bargelloni, and T. Patarnello.
  2002. Strategies for microsatellite isolation a
  review. Molecular Ecology 11 1-16.