Before Lab: The following paper is a made-up study, designed to illustrate some common methods and good and bad aspects of interpretation of phylogenetic studies. Read the paper before the lab that will meet during week 13. Use your textbook to make sure you can understand and explain all terms in bold face. NOTE: some information is in Chapter 10, which you should read thoroughly since it provides background on phylogenetic analysis in general. Other information can be found in Chapter 4 on mutation and Chapter 7 on molecular evolution. If there are terms on molecular evolution or mutation that are used in Chapter 10 and not defined, they're probably defined in Chapter 4 or Chapter 7. You can also use the glossary to help define terms, but you will frequently understand them better if you read them in context. If there are terms you do not understand, ask me, call me, or e-mail me BEFORE lab and ask questions! I will expect you to be able to explain these terms at the start of lab; I may call on specific students to define terms so be sure you can do so!

The study follows:

INTRODUCTION

Some plants have separate sexes (this is called being dioecious); in others, both sexes occur in the same plant (this is called being monoecious). A phylogenetic analysis is conducted of a previously undescribed group of plants, the yellowleafs, to study the evolution of these two forms of reproduction in plants.

METHODS

The phylogeny of six yellowleaf species, of which three are dioecious and three monoecious, was studied using a 250 base pair stretch of DNA from a protein coding gene from the choloroplasts (cpDNA; this is DNA that occurs in chloroplasts, not the nucleus; it is similar to mitochondrial DNA in that it is reproduced asexually, without recombination). The tree was rooted using three plant species that do not apparently fall within the yellowleaf group. Sequences were aligned by visual examination. An initial maximum parsimony analysis was made to determine phylogeny using a heuristic search which was initiated 3 different times using randomly chosen initial trees. Third positions changes, especially transitions, may evolve rapidly and be saturated.  Comparison of number of transitions and transversions between species pairs within yellowleafs and between yellowleafs and the outgroups indicated that third position transitions showed signs of saturation but other changes did not. Based on these results, a subsequent parsimony analysis of phylogeny was conducted in which third position transitions were not used. Transitions and transversions were weighted equally. A consistency index was calculated. Support for the different nodes in the tree was also evaluated using bootstrap analysis with 100 replicates. Another analysis of these data was done using maximum likelihood analysis. The evolution in monecy and dioecy was evaluated based on the resulting phylogeny.

RESULTS

Twenty base pair sites were determined to be phylogenetically informative and were used for the phylogenetic analysis. When all twenty sites, including transitions and transversions at all sites, were used, three equally parsimonious trees of cpDNA haplotypes from all species were obtained. When third position transitions are excluded from the analysis, a single most parsimonious tree was obtained. This does not differ greatly from any of the trees based on all sites, or from the tree obtained using maximum likelihood analysis, so this tree is used for our analysis of the evolution of monoecy and dioecy. A consistency index of 0.72 was obtained for this tree. Bootstrap support for the branches is given next to each branch in Figure 1.

Figure 1. Evolution of dioecy in the yellowleafs based on the most parsimonious tree excluding third position transitions; thick purple lines indicate the most likely evolutionary origins of dioecy. M: monoecious. D: dioecious.

CONCLUSIONS

The tree shows that monoecy is primitive to the yellowleafs; dioecy evolved in the ancestor to the red, blue, and spotted yellowleafs. The consistency index of 0.72 suggests only a moderate amount of homoplasy so the tree appears well supported; high bootstrap numbers for most branches also indicate good support for the tree.

Click here to return to the index of lectures

Click here to return to the introduction to the critique assignment