PLNT3140 INTRODUCTORY CYTOGENETICS October 2023

Karyotype Lab Assignment

This assignment is worth 30% of the lab grade.

Due date: November 10, 2023.

Goal: To measure the lengths and arm ratios of barley chromosomes

1. (10) Prepare slides of barley chromosomes using Fuelgen stain.

The procedure is described in KaryotypeLabManual.pdf. The lab instructor will demonstrated this procedure.

Once you have a good slide or slides, bring them to the photomicroscope to capture images, which you can then upload for further use.

2. (5) Learn how to measure chromosomes in the karyotype tutorial.

Create a folder called karyotype to hold all files related to this assignment. In the tutorial, you will create a subfolder within the karyotype folder called "example".

For this and other labs, we will use ImageJ, an image processing program produced by the NIH. ImageJ is already installed on the CCL Linux system. Alternatively, you can install ImageJ on your own computer and use it there.

Tutorial: Karyotypes with ImageJ

Save your cell image files, along with the following files into your karyotype folder:

karyotype.template.xlsx - spreadsheet for calculating arm length ratios and total chromosome lengths
karyotype.report.template.docx - template for karyotype report. Rename the file so that your filename will be unique. For example, replace "template" with your initials or last name.
ideogram.template.pptx - template for creating your ideogram

3. (5 points) Annotate an image of chromosomes for karyotype analysis

Go through your files to find an image with well-separated mitotic chromosomes. Ideally, chromosomes will not overlap, and be clearly visualized. Save a copy of this image in your karyotype folder.

Using the procedure described in the Karyotype tutorial, create an annotated karyotype of your chromosome image. This image will be inserted into your report, as seen in part 1 of karyotype.report.template.docx.

Hints:

Make sure to crop your image so that it is similar in size to the example. A full screenshot is not acceptible.
Save your final image in PNG or JPG format. Then, insert your image into the report under the appropriate subheading.

4. (5 points) Calculate chromosome lengths and arm ratios

Copy the results from your ImageJ Results window into karyotype.template.xlsx. As in the tutorial, only the arm lengths column should be copied to column B of the template. Save your spreadsheet. Next copy the spreadsheet into part 2 of the report.

5. (5 points) Create an ideogram based on the measured lengths of the chromosome arms

The report has an example of one way to draw an ideogram. Each chromosome is shown with the long arm above the dotted line, and the short arm below the dotted line. Your job is to create a similar ideogram that represents proportionally the relative lengths of the two arms for each chromosome. It is probably easiest to modify the ideogram in ideogram.template.pptx, but feel free to create your own drawing if you wish.

If you use the file provided, note that the lines and numbers for each chromosome are grouped. If you click on a chromosome, all parts will move as a single object. In order to change the lengths or numbers, you must select a chromosome, right click, and choose "Ungroup". Make your changes to the chromosome, and then select both arms and numbers, right click, and choose "Group". It is probably best not to ungroup more than 1 chromosome at a time.

Some things to note:

Chromosomes that can reasonably be assumed to be homologues should be placed together, and indicated as a pair using brackets or other markings. Ideally, homologous chromosomes will have similar total lengths, similar l and s arm lengths, and similar l/s ratios.
Chromosomes that cannot reasonably be matched in a pair should be well-separated (eg. 1-2 and 15-16)

Reference: NCBI Barley Genome Page

Submitting your assignment

Make sure your report includes your name, student number and UM email address.
This file contains places for pasting in data, as part of your report. It is NOT acceptable to upload your data as separate files. They must be inserted into the report.
Save your assignment report as a PDF file, and upload it to the PLNT3140 UMLearn dropbox site in the Karyotype Lab folder. Files in word processing formats (.doc, .docx, .rtf, .odt) are NOT acceptable.

If your don't understand how to do something, contact the teaching assistant.