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Section 6

EVOLUTIONARY MECHANISMS

INTRODUCTION

Definition
Evolution is a change that occurs over time in the genetic composition of a population.

More precisely:

Evolution is a change that occurs over time in the proportions of organisms in a population that differ genetically in one or more characteristics.

Question: What factors are capable of causing evolutionary change within a population?

Approach - Instead of attempting to conduct experiments with real creatures and real genes, we will first investigate these questions using a hypothetical population. We will conduct simulations to determine the factors that can facilitate or inhibit genetic change at one locus within this population. Our hypothetical population will be very simple, and we will focus on how different factors affect the genotype and allele frequencies at one locus.

The simulation approach we will use today represents a type of theoretical investigation. Why should we use this approach before we conduct experiments with real organisms? Theoretical inquiry serves as a guide for empirical research (i.e., research that involves taking measurements on real organisms). Real systems are complex, and experimental research with these systems is often time consuming and expensive. Theoretical research allows us to ask "what if" questions using very simple systems which we refer to as models. The results of such investigations can suggest questions that might prove profitable for empirical research and also can suggest types of results we might expect to obtain from such studies. If the predictions of theoretical investigations and empirical studies are at odds with each other, then we must create more complicated theoretical models to account for important factors that we omitted from our initial inquires. All active areas of research involve this type of interplay between theoretical and empirical research, and our understanding of how the world operates depends upon both types of investigations.

BEFORE YOU COME TO LAB

Read the synopsis of the Hardy Weinberg Equilibrium Theory on pages 28-31 of the lab manual and work the sample problems.

GAME PLAN

1. As a class we will simulate a very simple population of interbreeding organisms, and we will then investigate how changes in characteristics of the population and its environment affect the direction and rate of evolution at one particular locus.

2. We will then use a computer program to simulate a similar hypothetical population. The difference between the computer simulations and the ones we will conduct by hand is that the computer conducts the simulations faster than we can and it allows us to specify much larger population sizes than we can achieve with our small lab sections. The class simulations, during the first part of lab, should familiarize you with the rationale behind thecomputer simulations and make them be less of a "black-box" procedure.

3. Before we conduct each simulation, record your prediction for the simulation's outcome and the rationale you used to make that prediction.

4. At the end of the lab, I will assign each lab group data from one of the simulations. Before our next lab meeting, members of the group should meet and:

a. Discuss the outcome that each member predicted for the simulation and the reasoning behind these predictions.

b. Calculate genotype and allele frequencies for each generation.

c. Plot data from your assigned simulation and results predicted by the null model on two graphs (one for genotype frequencies, the other for allele frequencies) in a manner that will allow us to compare the results from your simulation with those predicted for a situation where no evolutionary mechanisms operate.

d. Prepare a group statement that compares and contrasts the results from your simulation with the predictions of the null model (i.e., Hardy Weinberg Equilibrium Theory). Provide your conclusions regarding the effect of the manipulation on evolution in our hypothetical population.

e. Elect a spokes-person to deliver your statement to the class at our next lab meeting.

5. Each student will complete the Evolutionary Mechanisms Problem Set and submit this assignment in lab next week. You have my permission to consult with other members of your lab group as you complete this assignment.

CLASS SIMULATIONS

Assumptions - Our goal is to simulate a very simple population and look at one very simple genetic characteristic. In order to accomplish this goal, we will assume that:

1. Each individual in the population reaches reproductive maturity, mates, produces two offspring, and then dies.

2. Individuals in the population are hermaphrodites (i.e., can function as both mothers and fathers) but cannot self-fertilize.

3. The genetic trait under consideration is controlled by two alleles, A and a, at one locus. The A allele is dominant with respect to the a allele. Individuals in the population are homozygous for all other loci.

4. No individuals enter or leave our population (i.e., no immigration or emigration).

General Instructions

1. Each new simulation will begin with a population with an initial frequency of 0.5 for alleles A and a, and genotype frequencies of 0.25 for the aa and AA genotypes and 0.50 for the Aa genotype.

2. Each of you will receive an envelope with two slips of paper that represent your genotype at the locus of interest (e.g., if you receive an envelope with two a slips, your genotype is aa). Record this genotype on your data sheet.

3. You will then proceed to mate. Unless instructed otherwise, you should be entirely promiscuous (remember, this is very safe sex - all on paper only). Choose anyone else in the class (male or female - remember for our simulation purposes you are all hermaphrodites) and confidently approach them. They will not refuse. Once you find a mate, flip a coin to determine which allele you will contribute to your first offspring - your mate will do the same (if you are homozygous you obviously can skip this step). Record the genotype of your offspring on a piece of scrap paper. Now, repeat the process to produce a second offspring and record it's genotype. By having each couple produce two and only two offspring per generation, we keep the population size constant.

4. Once you and your mate have produced two offspring, wait for me to signal the end of that generation. At the end of the generation, you and your mate will "die", and each of you will assume the genotype of one of your two offspring (e.g., if you produce an AA and an Aa offspring, one of you assumes the AA genotype and the other assumes the Aa genotype). Record your new genotype on your data sheet.

5. When I signal the beginning of a new mating session, you will pick another mate and produce two new offspring using the alleles from your new genotype.

6. We will conduct 5 generations of mating for each simulation.

IMPORTANT NOTES

Mate with only one individual each generation.

Do not move on to a new generation of mating until I instruct you to do so.

If you are heterozygous, you sample with replacement when you decide which allele to give to each offspring. This means that if you designate the A allele as heads and the other allele as tails, you could end up donating an A allele to both offspring.

Remember to record your new genotype in the appropriate place on your data sheet.

Simulations

Null Model

If no evolutionary mechanisms influence the locus under consideration, then allele frequencies should remain constant over time and genotype frequencies should eventually match Hardy Weinberg predictions. We will calculate the allele and genotype frequencies of our populations at the beginning of each simulation. If the allele and genotype frequencies we calculate at the end of each generation deviate significantly form the initial conditions, then we know that some evolutionary mechanism affected the population during the course of the simulation.

Simulation #1

In this simulation we will examine the effects of population fragmentation and small population size on evolution. I will randomly split the class into two populations of eight individuals each. Remember to clearly mark your data sheet with the name of your population. The individuals in each population will then proceed to mate following the instructions provided above. NOTE: Mate only with individuals in your population.

Simulation #2

Resume the genotype you were assigned at the beginning of simulation #1. In this case, we will again form one large population and examine the effects of non-random mating on evolution at our locus. We will again assume that the AA and Aa genotypes have the same phenotype (i.e., A dominant to a). When you pick a mate, keep trying to find one whose phenotype matches yours. Mate only with someone of a different phenotype as a last resort. Otherwise, proceed as you did for simulation #1.

EVOLUTIONARY MECHANISMS - DATA SHEET

SIMULATION # 1 MANIPULATION Population fragmentation

POPULATION Oranges

Initial Genotype

F1 Genotype

F2 Genotype

F3 Genotype

F4 Genotype

F5 Genotype

SIMULATION # 2 MANIPULATION Non-random Mating

Initial Genotype

F1 Genotype

F2 Genotype

F3 Genotype

F4 Genotype

F5 Genotype

COMPUTER SIMULATIONS AND PROBLEMS

Assumptions - Assume that coat color in a certain strain of mice is controlled by one gene with 2 alleles. One allele codes for black coats, and the other codes for white coats. In the population you find 3 coat phenotypes: black, gray (= heterozygotes), and white. The frequency of the black allele is 0.5 and the population starts out in Hardy-Weinberg equilibrium.

1. If you expect the observed genotype frequencies in the population to change substantially as the simulation proceeds (i.e., deviate substantially from those predicted by the Hardy-Weinberg equilibrium theory)
2. The nature of the change you expect (e.g., excess of both homozygotes and a deficiency of heterozygotes)
3. The ultimate outcome of the situation for the population (e.g., loss of the black allele from the population).

The Evolve Instruction sheets that follow provided detailed instructions for each simulation.

Simulations

A. A very large isolated population with no appreciable mutation in coat color alleles, random mating, and where individuals with white coats are spotted easily by predators but where mice with black and gray coats are less obvious to predators.

B. The same large, isolated population of mice with no appreciable mutation in coat color alleles, random mating, and where individuals with white coats are spotted the most frequently by predators, individuals with black coats are the next most frequently spotted, and gray individuals are rarely spotted by predators.

C. The same isolated population of mice, but with a small population size. Assume: random mating, no appreciable mutation in coat color alleles, and no differential survival among coat color phenotypes.

Bio 112 Evolve Instructions Peroni

To access Evolve software:
Turn on the computer (make sure it is plugged into the outlet). Double click on the hard drive icon, then double click on the Evolve folder icon. Double click on the Evolve icon.

Simulation A

Highlight the "Selection for Dominant Allele" line, then click on the "Start Problem" box.

Click on the "Change Parameters" box. Set the allele frequencies at 0.5, the number of generations the simulation will run at 50, and the population size at 8,000. Click on the "Genetic Drift" box and make sure that the maximum population size is set at 9,999 and the post-crash size is set at 8,000.

Let the dark circle represent the black allele and the open diamond represent the white allele. Set the "survival" of the homozygous black and heterozygotes at 30 and their "reproductive" output per year at 8. Set the survival and reproductive output per year of the homozygous white genotype at 22 and 5, respectively.

Click on "Update" then click on "Done".

Go to the Graphs menu located on the top menu bar and select either freq of the black allele or freq or heterozygotes (selected variables will have a check mark; to select a variable click on it; to insure that a variable with a check mark is not plotted, click on the variable name and the check mark will disappear).

Click on start, and the simulation will begin. Use the graphs menu to look at how the values of different variables changed during the simulation. Print out a copies of the most relevant graphs, and annotate them so you know the parameters used to generate each figure.

Now, modify the simulation to make the homozygous white genotype lethal at a point prior to reproductive maturity. To do this, click on the "Change Parameters" box. Set both the "survival" and "reproduction" of the homozygous white genotype to 0. Make sure all other parameters match the ones you used for the first simulation. Click on "Update", then click on "Done." Click on "New Trial" then click on "start." Print out a copies of the most relevant graphs.

Simulation B

Select the "File" menu from the menu bar and select "New Problem." When the screen prompts you, click on the "New" box. Highlight the "Selection for Dominant Allele" line, then click on the "Start Problem" box.

Proceed as you did with the first trial for simulation #1, with the following exceptions:

Set the "survival" and "reproduction" of the heterozygote at 30 and 8, respectively; for the homozygous black create settings of 25 and 5; and for the homozygous white create settings of 15 and 4. Print out a copies of the most relevant graphs and annotate them so you know the parameters used to generate each figure.

Click on the "Update" box, then click "done". Click "Start" to initiate the simulation. Examine changes in the frequency of the black allele, then hide this graph and examine changes in the frequency of the heterozygotes.

Repeat the simulation, but first change the parameters so that "survival" and "reproduction" equal 25 and 5, respectively for both of the homozygous genotypes. Print out a copies of the most relevant graphs and annotate them so you know the parameters used to generate each figure.

Simulation C

Select the "File" menu from the menu bar and select "New Problem." When the screen prompts you, click on the "New" box. Highlight the "Genetic Drift- Pop 80 - 100" line, then click on the "Start Problem" box.

Click on the "Change Parameters" box. Set the allele frequencies at 0.5, the number of generations the simulation will run at 50, and the population size at 80. Click on the "Genetic Drift" box and make sure that the maximum population size is set at 100 and the post-crash size is set at 80.

Click on the "Update" box, then click "done". Click "Start" to initiate the simulation. Examine changes in the frequency of the black allele. Print out a copies of the most relevant graphs and annotate them so you know the parameters used to generate each figure.

Click "New Trial" and then "Start" again. Repeat this process one more time, then compare the results of the three simulations.

Next, examine the effects of even smaller population sizes on the frequency of the black allele. Click on the "Change Parameters" box. Click on the "Genetic Drift" box and change the maximum population size to 50 and the post-crash size to 5. Change the "Initial Population Size" to 50. Click on the "Update" box, then click "done". Click "New Trial" and then "Start." Repeat this process two more times (total of 3 simulations with these parameters). Print out a copies of the most relevant graphs and annotate them so you know the parameters used to generate each figure.

ACKNOWLEDGEMENTS

The in class mating portion of this lab was adapted from one used in the General Biology Program at Duke University; its origin is attributed to Dr. Paulette Peckol (Smith College). Dr. Patricia Peroni developed the idea of using simulation programs to model the effects of different evolutionary mechanisms on the genetic composition of populations; Dr. Valerie Banschbach implemented this idea using the Bioquest Evolve software, and the write up included in this lab is Dr. Banschbach's. The synopsis of Hardy-Weinberg Equilibrium Theory and the Evolve Instructions section were written by Dr. Patricia Peroni.

EVOLUTIONARY MECHANISMS PROBLEM SET

Work the following problems at the end of lab or before you arrive at lab next week. All problems refer to the population of mice described for the computer simulations.

A. Assume that the frequency of the black allele in the population is 0.6, and that the population meets all of the expectations of the Hardy-Weinberg equilibrium theory. What are the expected genotype frequencies for the coat color locus in this population? Show your work.

B. If the population continues to meet the assumptions of the Hardy- Weinberg Equilibrium theory, do you expect the allele and genotype frequencies at this locus to change drastically over time? Explain your answer.

C. Now investigate the coat color locus in another large population of this mouse species. In a sample of 100 mice from this population you find 60 with black coats, 10 with gray coats, and 30 with white coats. Calculate the allele and genotype frequencies in this sample.

D. Is there evidence to suggest that evolutionary mechanisms are operating on the coat color locus in this population? (This will require you to compare your observed genotype frequencies with those predicted by Hardy Weinberg - show your work) If so, which evolutionary mechanisms would most likely cause such deviations from Hardy-Weinberg expectations?

CALCULATION OF ALLELE AND GENOTYPE FREQUENCIES &

HARDY WEINBERG EQUILIBRIUM THEORY

BACKGROUND

Population geneticists study frequencies of genotypes and alleles within populations rather than the ratios of phenotypes that Mendelian geneticists use. By comparing these frequencies with those predicted by null models that assume no operation of evolutionary mechanisms within populations, we can draw conclusions regarding the evolutionary forces that may influence individual populations. The Hardy Weinberg Equilibrium Theory serves as the basic null model for population genetics.

CALCULATION OF ALLELE AND GENOTYPE FREQUENCIES

Allele Frequencies

Consider an individual locus and a population of diploid individuals where two different alleles, A and a, can be found at that locus. If your population consists of 100 individuals, then that group possesses 200 alleles for this locus (100 individuals X 2 alleles at that locus per individual). The number of A alleles present in that population expressed as a fraction of all the alleles (A or a) at that locus represents the frequency of the A allele in the population.

To calculate allele frequencies for populations of diploid organisms:

1. Multiply the number of individuals in the population by 2 to obtain the total number of alleles at that locus.

2. Select one of the alleles for your first set of calculations. For example this example, we will choose the A allele from the example provided above.

Individuals homozygous for the A allele will each possess 2 A alleles. Multiply the number of A A homozygotes by 2 to calculate the number of A alleles that group possesses.

Heterozygotes will each possess only one A allele. The total number of A alleles in the population will equal:

(the number of A a heterozygotes) + (2 X the number of A A homozygotes)

3. The frequency of the A allele will equal:

total number of A alleles in the population
total number of alleles in population for locus

4. The frequency of the a allele will equal:

1 - frequency of the A allele

Genotype Frequencies

Consider the same population, locus, and alleles described above. Genotype frequencies represent the abundance of each genotype within a population as a fraction of the population size. In other words, the frequency of the AA genotype represents the fraction of the population that is homozygous for the A allele.

To calculate genotype frequencies for populations of diploid organisms:

1. Determine the number of individuals with each genotype present in the population. In the example used above, you would count the number of individuals with the following genotypes:

AA
Aa
aa

2. To determine the frequency of each genotype, divide the number of individuals with that genotype by the total number of individuals in the population.

Frequency of AA genotype = # AA individuals / population size

Frequency of Aa genotype = # Aa individuals / population size

Frequency of aa genotype = # aa individuals / population size

IMPORTANT NOTE:

UNLESS YOU KNOW THAT A POPULATION MEETS HARDY WEINBERG EQUILIBRIUM ASSUMPTIONS, YOU MUST USE THIS PROCEDURE TO CALCULATE GENOTYPE FREQUENCIES. IF YOU KNOW THAT A POPULATION MEETS HARDY WEINBERG EXPECTATIONS, THEN YOU CAN CALCULATE GENOTYPE FREQUENCIES USING ALLELE FREQUENCIES AND THE HARDY WEINBERG EQUATIONS (see below).

HARDY WEINBERG EQUILIBRIUM THEORY

Assertions of the Theory

The Hardy Weinberg Equilibrium Theory refers to loci within populations that experience no evolutionary mechanisms (i.e., forces). For such populations the theory asserts that:

1. Allele and genotype frequencies should remain constant from one generation to the next. That is, no evolution should occur at these loci.

2. Given a certain set of allele frequencies, genotype frequencies should conform to those calculated using basic probability. In a one locus/ two allele system such as the one described above, the genotype frequencies should be as follows:

Frequency of AA genotype = (frequency of A allele)2
Frequency of aa genotype = (frequency of a allele)2

Frequency of Aa genotype = 2 X (frequency of A allele) X (frequency of a allele)

If the genotype frequencies obtained from a real population do not agree with those predicted by the Hardy Weinberg Theory, then population geneticists know that some evolutionary mechanism or mechanisms must operate on the locus of interest. A knowledge of the theory can help them narrow down the possible mechanisms. Then, they can use experiments to determine which potential mechanism or mechanisms operate on the locus. As such, the Hardy Weinberg Equilibrium Theory serves as an important tool for population geneticists.

Assumptions of the Theory (Evolutionary Mechanisms)

Populations will conform to the Hardy Weinberg Theory assertions only if no evolutionary forces or mechanisms influence the loci under consideration. The assumptions that populations must meet in order for the Hardy Weinberg assertions to hold include:

1. Large population size (i.e., no genetic drift)

2. Random mating

3. No difference in the mutation rates between alleles at the same locus

4. Reproductive isolation from other populations (i.e., no gene flow)

5. No differential survival or reproduction among phenotypes (i.e., no natural selection)

Example

Consider a population of 1000 individuals and the locus and alleles described above. Assume that you have no information on the presence or absence of evolutionary mechanism in this population. You find that the population consists of:

90 individuals homozygous for the A allele (AA genotype)

490 individuals homozygous for the a allele (aa genotype)

420 heterozygotes (Aa genotype)

1. Calculate the genotype and allele frequencies for this locus.

2. Determine if this population meets Hardy Weinberg Assumptions (in other words determine if evolutionary mechanisms operate in this population).

Calculation Allele and Genotype Frequencies

Since you do not know if this population meets Hardy Weinberg Assumptions, you must calculate both the allele and genotype frequencies using the raw data.

Allele Frequencies:
The frequency of the A allele will equal:

total number of A alleles in the population = [(90*2) + 420] = 0.30
total number of alleles in population for locus (1000*2)

The frequency of the a allele will equal:

1 - 0.03 or:

total number of a alleles in the population = [(490*2) + 420] = 0.70
total number of alleles in population for locus (1000*2)

Genotype frequencies:

Frequency of AA genotype = # AA individuals / population size = 90/1000 = 0.09

Frequency of Aa genotype = # Aa individuals / population size = 420/1000 = 0.42

Frequency of aa genotype = # aa individuals / population size = 490/1000 = 0.49

Hardy Weinberg Predictions

If no evolutionary mechanisms operate on this locus, then the Hardy Weinberg Equilibrium Theory predicts that the genotype frequencies should be as follows:

Frequency of AA genotype = (frequency of A allele)2 = (0.3)2 = 0.09

Frequency of aa genotype = (frequency of a allele)2 = (0.7)2 = 0.49

Frequency of Aa genotype = 2 X (frequency of A allele) X (frequency of a allele)
= 2*0.3*0.7 = 0.42

Conclusion

Since the observed genotype frequencies equal those predicted by the Hardy Weinberg Equilibrium Theory, we conclude that no evolutionary mechanisms operate on this locus in this population (i.e., the population meets the assumptions of the Hardy Weinberg Theory).

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