1. Phylogenetics and Coalescence Lab 9 October 24, 2012

2. Goals Construct phylogenetic trees using the UPGMA method Use nucleotide sequences to construct phylogenetic trees using UPGMA, NJ, and Maximum Parsimony methods Use coalescent simulation to determine historical change in N e Interpret coalescent trees to draw inferences about human migrations

3. Phylogenetic Methods Scope of the problem – Number of possible unrooted trees for n OTUs: – For 10 taxa -> 2,027,025 possible unrooted trees. – Need an optimality criterion

4. Phylogenetic methods A.Distance methods. 1. Unweighted Pair Group Methods using Arithmetic averages(UPGMA). 2. Neighbor Joining (NJ). 3. Minimum evolution(ME). B. Character based methods. 1. Maximum Parsimony (MP). 2. Maximum Likelihood (ML). 3. Bayesian Method (BA)

5. UPGMA Taxa1234567 HumanTGCGTAT ChimpanzeeTGGGTAT GorillaTGCGCTT OrangutanTGCTGTG GibbonTAGTAGC Step 1: Generate data (Sequence/ Genotype/ Morphological) for each OTU.

6. Distance can be calculated by using different substitution models : 1. # of nucleotide differences. 2. p-distance. 3. JC distance 4. K2P distance. 5. F81 6. HKY85 7.GTR etc Step 2: Calculate p- distance for all pairs of taxa. Taxa123*4567 HumanTGCGTAT ChimpanzeeTGGGTAT = 0.142857143

7. Step 3: Calculate distance matrix for all pairs of taxa and select pair of taxa with minimum distance as new OTU. TaxaHuChGoOrGi Hu0 Ch0.14280 Go0.28570.42850 Or0.57140.71420.42850 Gi0.85710.71420.85710.71420 Human Chimpanzee 0.714

8. Step 4: Recalculate new distance matrix, assuming human and chimpanzee as one OTU. taxaHu+chGoOrGi Hu+Ch Go Or Gi = 0.3571 taxaHu+chGoOrGi Hu+Ch0 Go0.357140 Or0.642850.42850 Gi0.785710.85710.71420 TaxaHuChGoOrGi Hu0 Ch0.14280 Go0.28570.42850 Or0.57140.71420.42850 Gi0.85710.71420.85710.71420

9. Step 5: Select pair of taxa with minimum distance as new OTU. Human Chimpanzee 0.071 Gorilla 0.179 0.107

10. Step 6: Again select pair of OTU with minimum distance as new OTU and recalculate distance matrix. taxa(Hu+ch)GoOrGi (Hu+ch)Go Or Gi = 0.5714 taxa(Hu+ch)GoOrGi (Hu+ch)Go0 Or0.57140 Gi0.80950.71420 TaxaHuChGoOrGi Hu0 Ch0.14280 Go0.28570.42850 Or0.57140.71420.42850 Gi0.85710.71420.85710.71420

11. Step 7: Again select pair of taxa with minimum distance as new OTU. Chimpanzee Human 0.071 Gorilla 0.179 0.107 Orangutan 0.286 0.107

12. Step 8: Again select pair of OTU with minimum distance as new OTU and recalculate distance matrix. taxa((Hu+ch)Go)OrGi ((Hu+ch)Go)Or Gi = 0.7857 taxa((Hu+ch)Go)OrGi ((Hu+ch)Go)Or0 Gi0.78570 TaxaHuChGoOrGi Hu0 Ch0.14280 Go0.28570.42850 Or0.57140.71420.42850 Gi0.85710.71420.85710.71420

13. Step 9: Again select pair of OTU with minimum distance as new OTU and make final rooted tree. Chimpanzee Human 0.071 Gorilla 0.179 0.107 Orangutan 0.286 0.107 Gibbon 0.393 0.107

14. Branch Supports 1.Bootstrap support. 2.Jack-knife support. 3.Bremer support. 4.Posterior probability support.

15. Bootstrap support Step 1: Randomly make “n” pseudo-replicates of the data with replacement and make tree from each replicate. Taxa2234677 HumanGGCGATT ChimpanzeeGGGGATT GorillaGGCGTTT Taxa1356724 HumanTCTATGG ChimpanzeeTGTATGG GorillaTCCTTGG Taxa1234567 HumanTGCGTAT ChimpanzeeTGGGTAT GorillaTGCGCTT

16. Bootstrap support Step 2: Make consensus tree of trees obtained from all pseudo replicates.

17. Phylogenetic Software available 1.PAUP. 2.Phyllip. 3.MrBayes. 4.Mega.

18. Problem 1. File mt_primates.meg contains the sequence data used to calculate the genetic distances in Example 1. Use Mega to build phylogenetic trees based on: 1.UPGMA. 2.The NJ Method. 3.Maximum Parsimony. Compute bootstrap confidence in the internal nodes of each tree. Compare the trees derived using each of these methods. Which do you think is the most informative? Does the computational efficiency of the UPGMA method result in misleading results in this case?

19. Problem 2. File pdha1_human.meg contains haplotypes detected by sequencing a 4.2-kb region of the X-linked Pyruvate Dehydrogenease E1 α Subunit (PDHA1) in 16 African and 19 non-African males. Use Mega to build a phylogenetic tree based on the NJ Method and interpret the results in the light of hypotheses about the origin of modern humans (see Example 11.4, p. 620-621, as well as p. 618 in Hedrick 2005).

20. Coalescence Wright-Fisher Model Until now we have implicitly used the Wright- Fisher Model Computationally expensive

21. Wright Fisher

22. The Discrete Coalescent Probability that two genes have MRCA j generations ago Probaility that 2 genes out of k have a common ancestor j generations ago Probability of no coalescence for j – 1 generations Probability of coalescence in the jth generation Probability of no coalescence in k lineages for j – 1 generations Probability of coalescence in the jth generation

23. The Continuous Coalescent Can derive continuous exponential function from discrete geometric representation Waiting time (T) for k genes to have k-1 ancestors (See math box 3.2 in Hamilton, 2009)

24. The Continuous Coalescent Model of population growth underlies coalescence Exponential Growth Bottleneck Hein et al. 2005

25. Coalescent Applications Coalescent topologies can be dependent upon convolution of N e and μ, migration rate, selection, recombination rate. Applications – Estimating recombination rates – Estimating historical migration rates between poulations – Estimating tMRCA – Estimating historical effective population size – Estimating strength of selection

26. From Data to coalescence Suppose we observe n genes with k mutations We want to get θ=4Neμ but do not know its true value Can calculate likelihood of θ for a bunch of possible values and find the one with highest probability

27. MCMC 1.Sample a new history from a distribution of histories (topologies + waiting times) 2.Divide the likelihood of this new history by the likelihood of the last history sampled 3.With probability proportional to this likelihood ratio, move to the new point. 4.Repeat steps 1-4.

28. Problem Fossil and molecular based evidence have both provided strong evidence for the divergence of the human and chimpanzee lineages approximately 6 MYA. However, timings and locations of human expansions beyond Africa have proved controversial. Use the Bayesian MCMC software BEAST to derive coalescent trees for sequences from the X-linked Pyruvate Dehydrogenease E1-alpha subunit gene that you also analyzed in Problem 2