Rebecca Schaeffer and Dr. Keith Crandall, Integrative Biology

Introduction

Many species of plants in the Leguminosae family are important economically, but many of these plants are now considered rare, threatened, or endangered. Seventy percent of the species in the genus Pediomelum are highly limited in geographic distribution, which influences conservation status. In addition to this, very little work with phylogenetic studies has been done on this group. Learning about evolutionary relationships within these species is a first step to understanding more about their conservation status.

Molecular phylogenetic studies provide a way to determine possible evolutionary relationships, developing something similar to a family tree for different species of organisms by comparing the sequences of gene regions in their DNA. Computer programs can be used to analyze DNA from different species and estimate evolutionary relationships in the form of a phylogeny, or family tree. This can tell us how closely each species is related to each other, how recently species diverged from each other, and how genetically different the species are. This information can then be used in a variety of ways, including providing knowledge that can be combined with other research to aid in developing conservation strategies. This project collected DNA sequence data from trnL/F and trnS/G chloroplast regions, performed phylogenetic analysis on those regions, and briefly discussed relevance to conservation status of the species.

Methods

First, plants were collected from all over North America by Ashley Egan, and DNA was extracted from dried leaf material. Two chloroplast DNA regions, trnL/F and trnS/G, were used in this project. The polymerase chain reaction (PCR) was used to amplify the DNA region. The PCR products were then visualized using gel electrophoresis to make sure the DNA amplified. DNA from a species that we knew worked properly was included as a positive control and a negative control with no DNA was included as well. This way, we could check the accuracy of our results. PCR products were purified and sequenced. Then, sequences were aligned using the program MUSCLE. Phylogenies were then estimated using maximum parsimony in the program PAUP.

The most common setback that occurred during this research was that a PCR reaction wouldn’t work properly and I’d have to figure out what went wrong. Sometimes the cause was just a missing reagent or that the amount of a reagent had to be changed. At one point, the distilled water that was being used had to be changed because there was some contamination in the water. For the most part, though, things ran smoothly and I think the biggest challenge for me was just learning how to use the computer programs.

Results and Discussion

The results from this project are shown in the phylogenetic trees in figure 1. The more closely two species are related, the more closely their branches will meet on the tree. As figure 1 illustrates, most species were grouped by genera, except for Psoralidium. Outgroups were used to root the tree and are the distantly related species found at the bottom of the tree. A large group of Pediomelum is unresolved, shown by multiple branches coming off from the same point. This shows a lack of genetic variation within this genus, which could be due to various causes, such as gene flow, cross pollination, young or recent origin, or recent, rapid speciation.

The young status of Pediomelum implied by the phylogeny means that speciation is still occurring, which has implications for conservation management. Ongoing speciation indicates that extinction rates may be high or there may be a small range size, since they haven’t had much time to disperse. Conservation measures may need to be taken to safeguard these species.

Conclusion

This project contributed to understanding evolutionary relationships in Psoraleae tribe of the Legume family. In addition, we now know that Pediomelum lacks genetic variation and may represent a recent, rapid radiation. More DNA regions are needed to increase the resolution of this group in order to really understand the evolution of this group. This is being addressed by incorporating this project into a larger project represented by Ashley Egan’s dissertation.

This project was a great opportunity for me to learn more about scientific processes and methods as well as understand DNA and evolutionary relationships better. This understanding will help me be a more effective science teacher because I’ll be able to share with my students my personal experiences with the scientific method and laboratory experiences. Performing this research has also shown me the importance of giving my students hands-on laboratory experiences to teach them science and to help them understand the scientific method.