Biology classes often include a lesson on human characters to illustrate basic Mendelian genetics. Most of you at some point have been asked whether you can roll your tongue, whether your earlobes are attached, or whether you have a hitchhiker's thumb. Unfortunately, the textbooks are wrong; most of the human traits used to illustrate genetics are either not genetic at all, or the genetics are more complicated than the simple one-locus, two-allele model. Plus, if these easily visible characters really did fit a simple genetic model, some students would find out from a classroom exercise that the person they call "Dad" isn't their biological father, and that would be awkward. So it would be nice to have easily observable characters in some other familiar organism that could be used to illustrate genetic principles.

In this lab, you will look at four characters in domestic cats, *Felis catus* (also sometimes called *Felis domesticus, Felis sylvestris catus,* or *Felis sylvestris domesticus*). You will collect data by looking at pictures of cats up for adoption in animal shelters. For the *orange* locus, it is well-established that there is one sex-linked locus with two alleles. You will be able to calculate allele frequencies and compare the genotype frequencies in females to those expected under Hardy-Weinberg equilibrium. For two loci, *spotting* and *longhair*, it is not entirely clear how well they fit a simple genetic model. You will collect some data on these two loci, analyze it as if they do fit the simple genetic model, then think about how you could investigate them in more detail. The *white* locus fits a simple genetic model well, with white dominant over non-white. Finally, you will think about possible problems with your sample and better ways of obtaining a more random sample of cats.

The goals of this lab are to reinforce basic concepts in Mendelian genetics (dominant/recessive, codominance, locus, allele), population genetics (Hardy-Weinberg), population sampling, and statistics. Specific questions to be addressed are whether genotype frequencies at the *orange* locus fit Hardy-Weinberg proportions, and whether allele frequencies are correlated with climate.

For those of you who will become teachers, variations on this lab are a good introduction to genetics for students ranging from third grade ("Are there more long-haired cats in colder areas?") to grad school ("What is the molecular basis of variable expression of piebald spotting?").

I will assign you one large city in the US or Canada. You will be responsible for collecting data on 100 cats from that city, 50 female and 50 male. You will record the information in a spreadsheet; download one spreadsheet for females and one spreadsheet for males.

First, go to Petfinder.com. Enter your city and state under "Location," choose "Cat" under "Type," choose "Adult" and "Senior" under "Age," and choose "Female" under "Gender." Then click "Search." You should get a list of cats for adoption in and near the city you entered. Click on each thumbnail picture to get more details on each cat. There may be more than one picture of each cat, and clicking on the pictures may enlarge them.

Look at the pictures of the first cat. Does one of the pictures show one entire front leg, from the paw to the shoulder? If no, go on to the next cat. If one picture does show the entire front leg, record the following information in the spreadsheet:

**Name:** So you don't score the same cat twice, and because some people give their cats goofy names.

**Hair length:** long or short. Hair length is said to be controlled by the *longhair* locus, with the alleles L and l. ll cats have long hair, while Ll and LL cats have short hair. Are there really just two hair lengths, or are there cats with medium-length hair? If a picture or description makes it seem like a cat has medium-length hair, take careful notes; dealing with this kind of ambiguity is an important part of science, don't just sweep it aside.

**White or non-white: **The *white* locus has two alleles. W is dominant, so cats with the WW or Ww genotypes have completely white fur. Cats with the ww genotype have some non-white fur. Note that cats with the SS genotype at the *spotting* locus can be mostly white, but will have at least some colored hair, often on the top of their head or their tail.

**Orange/cream color present or not: **The *orange* locus is on the X chromosome, so males are either OY or oY. An OY male is orange or cream colored, while an oY male is black, brown or gray. The darkness of the color (orange vs. cream or black vs. brown vs. gray) is determined by other genes that we won't try to score. An OO female is orange or cream colored, while an oo female is black, brown or gray. In an Oo female, one allele is inactivated in each cell early in development. Cells descended from a cell in which the O allele was inactivated will make black fur, while cells with the o allele inactivated will make orange fur. The result is a cat with patches of orange and black (or cream and gray) fur. This is generally called a "tortoiseshell" if there are no white patches and a "calico" if there are white patches; we'll call them all "calico" for simplicity.

**Presence and amount of white spotting:** Record whether each cat has some white patches on it, or is completely colored. Cats with the *ss* genotype at the *spotting* locus have no white fur, while the *Ss* and *SS* genotypes have white patches. The extent of white fur in *Ss* and *SS* cats is variable, being determined by other genes and by environmental factors. Some sources say that cats with the *SS* genotype have white fur on more than half their body, while cats with the *Ss* genotype have white on less than half their body. One of the main goals of this project will be to collect a large number of pictures of cats, and see if the ones with white spotting divide neatly into two categories, "less than 50% white" or "more than 50% white," or if there's a continuous range of amount of white fur.

Copy the best picture of the cat (control-click on a web picture to copy it) and paste it into the spreadsheet. Shrink the picture to fit the column width.

Count the number of long-haired cats and short-haired cats separately for males and females. Compare the proportion of long-haired cats between males and females using a chi-square test of independence. Then combine the male and female data and estimate the frequency of the *l* allele using the Hardy-Weinberg relationship. Write your city, your name, and the allele frequency on the board, so we can compare the frequencies in different cities and see if there's a correlation with climate.

Count the number of all-white cats and non-all-white cats separately for males and females. Compare the proportion of white cats between males and females using a chi-square test of independence. Then combine the male and female data and estimate the frequency of the *w* allele using the Hardy-Weinberg relationship. Write your city, your name, and the allele frequency on the board, so we can compare the frequencies in different cities and see if there's a correlation with climate.

For the Orange locus, you don't need to estimate the allele frequencies using Hardy-Weinberg, you can count the alleles directly. First, count the number of orange males and black males (remember that "orange" includes cream, and "black" includes gray and brown). Omit the all-white cats. Each orange male has one *O* allele, and each black male has one *o* allele.

Next, count the orange females, the black females, and the calico or tortoiseshell females. Each orange female has two *O* alleles, each black female has two *o* alleles, and each calico or tortoiseshell female has one *O* and one *o* allele.

Use the chi-square test of independence to compare the proportions of black and orange alleles between males and females from your city. Then combine the data and calculate the overall allele frequency.

Calculate the expected proportions of orange, black, and calico cats using the Hardy Weinberg relationship. If the allele frequencies in males and females were not significantly different, use the allele frequency based on the combined data; if the frequencies were significantly different between males and females, just use the female allele frequency. Test the fit to Hardy-Weinberg using the chi-square test of goodness-of-fit. In the spreadsheet, enter 1 for "degrees of freedom (intrinsic hypothesis)."

Many sources say that you can tell *SS* cats from *Ss* cats because *SS* cats have more than 50% of their body covered in white fur, while *Ss* cats have less than 50% white fur. I don't know whether there's good evidence for that, or even if cats with some white fur can be neatly divided into two categories. To investigate this, add a row to your cat spreadsheet labelled "percent white." Look at each cat picture, and give it an estimated percentage of white fur, from 0 (for the cats with no white fur) to somewhere in the 90s. Ignore the all-white cats.

Once you have given a number to each cat, save your spreadsheet, then sort the columns based on the percentage white. Adjust the numbers and sort the cats again. You want the cats to be sorted from least white to most white, in your subjective estimate of the percentage white.

After you've sorted the cats from least to most white, look at their pictures and see how you would group the ones with some white fur into two categories, "less white" and "more white." Write down the boundary cats; for example, your boundary might be between Ms. Bootsykin and JoJo. Don't tell anyone where you think the boundary is.

Now hide the row that has your estimate of the percentage white. Ask four of your classmates, one at a time, to look at your cat pictures and decide where the boundary is between "less white" and "more white." Have them write down the boundary cats, but don't let them discuss their choice with anyone. Once you and your four classmates have made their choices, see whether everyone put the boundary at the same place.

Using your boundary between "less white" and "more white" and assuming that "less white" are *Ss* and "more white" are *Ss*, calculate the allele frequencies and test to see whether the genotype frequencies fit the Hardy-Weinberg expectations.

You must turn in a written report on your cat results; I'll tell you the due date in class. It must contain the following information:

- A
*short*description of what you did, including the name of your city, and how you resolved any ambiguous phenotypes (for example, if a cat was described as "long haired" but looked short haired in the picture, what did you do?). You don't need to repeat everything from the instructions, just what you did that might be different from what other students did. - A table with the numbers of cats of each phenotype, separated by sex, for the long hair, white hair, and orange loci. The table should also include the results of your statistical tests comparing the phenotype frequencies (for long hair and white hair) or allele frequencies (for orange) between males and females. The table should include the allele frequencies you estimated for long hair and white hair using the Hardy-Weinberg relationship. For each locus where the phenotype or allele frequencies were not significantly different between males and females, include the phenotype counts and estimated (white hair and long hair) or counted (for orange) allele frequencies for males and females combined.
- A few sentences about the test of fit to Hardy-Weinberg proportions of the orange locus in females. Include the P-value and whether there are more or fewer heterozygotes than expected. If you got a significant deviation from Hardy-Weinberg proportions, write a few sentences speculating about why you might have gotten that result, and describing an additional experiment to test your speculation. Your additional experiment must be something other than just collecting a larger sample of cats.
- A table with your results on the white spotting locus. You should have one line for you and one for each of your four classmates or friends. Each line should have the number of cats with no white (this should be the same for everyone), the number of cats with "less white," and the number of cats with "more white." It should also include the names of the "boundary cats," the ones that each person decided marked the boundary between "less white" and "more white." The table should also include the allele frequencies and the P-value for the test of fit to Hardy-Weinberg proportions for each person, assuming that "less white" are heterozygotes and "more white" are homozygotes. Of course, if everyone picked the same "boundary cats," the numbers will be the same for everyone; if they picked different "boundary cats," then numbers will be different.

You must also e-mail your spreadsheet of raw data (including the cat pictures) to me at mcdonald@udel.edu.

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This page was last revised August 28, 2017. Its URL is http://udel.edu/~mcdonald/geneticslab1.html