1997 WWLPT Biology Institute:  Life Cycles:  Reproduction & Embryological Development

 

by   Carol Bernon     BARNSTABLE HIGH SCHOOL    744 WEST MAIN STREET      HYANNIS, MA.  02563              gbernon@capecod .net

 

Teacher demonstration or group/cooperative activity.

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• Biology and A.P. Biology students
• Grades 9 -12

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Notes to the Teacher: to top
The major focus of this activity is for students to see embryogenesis in action.  If eggs can be found in early stages of  development ( 1 - 8 cells, for example) cleavage can be witnessed and nuclei can be observed  to divide in as little as 10 -15 minutes.  For interested students and teachers, this can be a very exciting and inspiring experience.

For group work, students must be able to use dissecting and compound microscopes.  Cultures should be ordered at least 2 weeks in advance from the Caenorhabditis Genetics Center from the University of Minnesota (see references) or from local research centers or universities that use C. elegans.    Worms should be kept at  20 - 22 degrees Celsius, and temperatures above 25   result in sterility.

This activity may be done as a demonstration or group lab.  For demonstrations,  one slide may be set up on a demonstration microscope and displayed on the video monitor.  Embryogenesis may  be viewed for several minutes or several hours, and may extend for several days, if additional media is available, so that development into the adult form can be observed.  

If students are working individually or in groups,  appropriate precautions should be taken when using single-edged razor blades.  No special problems are presented with the use C. elegans or the medium in which it is cultured.
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         Cultures of Caenorhabditis elegans wild type (N2 strain)
         C. elegans media
         Compound and dissecting microscopes
         Video camera attachment for microscope and  monitor screen*
         Single-edged razor blades
         Worm picks
         Slides and coverslips
         Pipettes
 
*For demonstrations.

Hermaphrodites carry out self-fertilization, or, preferentially, are fertilized by males. Over 300 eggs are produced by a single hermaphrodite, and embryogenesis and may be viewed in various stages while still in the body cavity of the parent.   The fertilized egg undergoes the first cleavage within 40 minutes, in which the anterior and posterior regions are formed.  Successive cell divisions may occur at intervals of 15 minutes or less.  Transparent bodies and eggs allow structures to be easily seen when viewed with a light microscope under high power.  Micrographs showing the various stages of embrogenesis (Schierenberg, 1986) are available at Indiana University.

Worms are available on a culture dish of medium with a lawn of E. coli, the primary food for the nematode.  Individual worms may be transferred by gently picking them up with a "worm pick" made by melting fine platimum wire on the end of a pasteur pipette.  Another method of harvesting many worms is to flood the culture dish with water and transfer the worm suspension to a petri plate.   Using the dissecting microscope, hermaphrodites may  be selected and trasferred on to a small drop of water on a glass slide.

The eggs are more easily studied after removal from the parent body.  Using the dissecting microscope, locate the hermaprodites.  With a new single-edged razor or scalpel blade, cut the worm into two or more pieces.  The internal pressure will force eggs out.  Cover with a cover slip and examine under the compound microscope.

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        PROBLEM:
 
        How do organisms develop from a fertilized egg?
        How long does it take for a cell to divide?
        How does a zygote develop into an embryo?

        Observe the stages of development of the eggs of C. elegans, a common
                    soil nematode.

        MATERIALS:
 

         Culture of nematodes:  Caenorhabditis elegans
         Compound and dissecting microscopes
         Single-edged razor blades or scalpels
         Worm picks
         Slides and coverslips
         Pipettes
         Lab notebook

PROCEDURE:

Part I :  Getting to know your worms.

Obtain a culture of C. elegans and examine under the dissecting microscope.  Notice the movement of the worms over the media and try to determine the anterior and posterior ends from the direction of movement.  Carefully, using the pick, transfer some worms from the culture dish to a drop of water on a slide and place a cover slip over the specimen.  Examine the slide under the compound microscope, first under low power, then under increased magnification.  Watch the behavior of worms and identify hermaphrodites and males.

Part II.  Examination and study of eggs.

Using the dissecting microscope, select 3 or 4 hermaphrodites from your cultures.  Transfer them to a small drop of water on a glass slide.  Place the slide uncovered on the stage of the dissecting microscope.  While looking through the microscope, locate your worms.  With a new single-edged razor or scalpel blade, carefully cut the worms into two or more pieces.  The internal pressure will force eggs out.  When you have finished cutting the worms, add a cover slip and tranfer the slide to the compound microscope.  Examine them first under low power to locate individual eggs, then increase magnification.

Locate an egg and determine the number of cells it contains.  Try to find one with a very few visible nuclei.  Determine what other cellular structures are visible.  If a 2-celled stage has been located, one cell should be larger than the other.   The larger cell will divide first to form the anterior region of the worm.

Make a sketch of your egg and note the time.  Record any details.  Check your eggs every few minutes and look for changes.  Record them and make additional drawings.  Compare your egg with those of other students.  

When the period is over, save your eggs for later observation or clean up your materials as directed.
 

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For Films and Cultures of C. elegans:

Caenorhabditis Genetics Center
Gortner Avenue
University of Minnesota
St. Paul, MN 55108.
attention: Theresa Stiernagle

References

Riddle D.L, Blumenthal, T., Meyer, B.J. and Priess, J.R.  1997.  C. elegans II.
Cold Spring Harbor Laboratory Press.

Wood, W.B. and the Community of C. elegans researchers. 1998.  The nematode Caenorhabditis elegans.   Cold Spring Harbor Laboratory Press.
 

Films

Embryonic Development of the Nematode C. elegans.  Einhard Schienberg/IWF.  12 minutes.

A Nematode as a Model Organism.  The Genetics of Programmed Cell Death. 30 minutes.
H. Robert Horvitz, Howard Hughes Med. Inst.  MIT.  1966.  Cognito Learning Media Inc.

Website

http://eatworms.swmed.edu

sunflower.bio.indiana.edu