Introduction: (Initial Observation)
Certain invertebrate aquatic animals are able to regenerate their missing parts. This regeneration ability in some animals is so strong that if you cut the animal in half, each half will grow to a complete a whole new animal. The tail half regenerates a new head and the head half regenerates a new tail.
This unusual ability also is a method of asexual reproduction for such animals.
Regeneration abilities vary among these animals; however, they are all subject of attention by scientists who are trying to identify the genes responsible for such regeneration.
The purpose of this project is to observe the regeneration ability of these animals. I may either select sponges or planarians for my experiments because these two are visible to the naked eye and I do not have to use microscope and complex tools for my observations. I may also try to study the effect of one environmental factor such as temperature or pH on the rate of regeneration.
Information Gathering:
Find out invertebrate aquatic animals and their reproduction methods. Read books, magazines or ask professionals who might know in order to learn about regeneration in plants and animals. Also gather information about the specific animal that you want to study (sponge, planarian,…). Keep track of where you got your information from.
Following are samples of information that you may gather.
What is regeneration?
Regeneration is replacement of parts that have been cut off or otherwise removed:
By growth and division of cells, often dedifferentiation & redifferentiation, & also cell rearrangements.
Some kinds of animals can regenerate all parts of their bodies: sponges, Hydra, Planaria, and some sea squirts. Note that all these also undergo asexual reproduction=budding of new individuals from the body of another. Most or all starfish can regenerate arms from the body; but only those of one genus (Linckia) can regenerate the whole body from arms! (contrary to what most people think) These deliberately tear arms off as a means of asexual reproduction.
Sponges:
Sponges are the simplest form of multi-cellular animals. A sponge is a bottom-dwelling creature which attaches itself to something solid in a place where it can find enough food to grow. The scientific name for sponges is “Porifera,” which translates into “pore-bearing.”
Almost all sponges pump water to obtain nutrients and oxygen. A sponge is covered with tiny pores which lead internally to a system of canals and eventually out to one or more larger holes. These canals exist to move water through the sponge’s body. Lining these canals are special collar cells. The collar cells force water through the sponge which brings oxygen and nutrients while removing carbon dioxide and waste. The collar cells are how sponges feed. The water brings with it bacteria and other organisms which the cells capture and filter out.
How Do Sponges Reproduce?
Most sponges are both male and female. During mating, one sponge plays the male role while the other plays the female role, even though both are capable of playing either role. A sponge may play a female role one time and a male role the next time. Sperm is released by the “male” sponge and enters the “female” sponge. After internal fertilization, larvae is released. After floating around for a few days, they settle down and start growing.
Sponges can regenerate the entire organism from just a conglomeration of their cells.
Planarians:
These flatworms can regenerate the entire body from just a small piece. They do this by the proliferation and differentiation of the totipotent stem cells that it retains in its body throughout its life.
Paramecia:
Paramecia can not regenerate missing parts like Planarians and sponges; however they also have some unusual properties or abilities that I like to call degeneration.
In paramecia, if you (somehow) rotate a small area of the animal’s surface so that the power strokes of the cilia in this area are pointing backward, then providing that this area is about half-way between the anterior and posterior ends (which is the plane in which these animals divide after mitosis), then both offspring will have areas of reversed cilia; and this will be inherited by all their offspring, without limit.
If you fuse two Paramecia, back to back, so that they have two mouths on opposite sides (Janus morphology) then this double body will be inherited by both daughter cells each time mitotic divisions occur.
Question/ Purpose:
What do you want to find out? Write a statement that describes what you want to do. Use your observations and questions to write the statement.
The purpose of this project is to experiment and observe the regeneration process in planarians or sponges. Record your daily observation so you can use it to answer questions like:
- What is the rate of regeneration in planarians?
You can also perform additional experiments to determine the effect of environmental factors on the regeneration of planarians. Sample questions are:
- How does pH affect the rate of regeneration in planarians?
- How does light affect the rate of regeneration in planarians?
- How does temperature affect the rate of regeneration in planarians?
Identify Variables:
When you think you know what variables may be involved, think about ways to change one at a time. If you change more than one at a time, you will not know what variable is causing your observation. Sometimes variables are linked and work together to cause something. At first, try to choose variables that you think act independently of each other.
For the first question, we need to determine how long it takes for a fragment of planarian to regenerate and become a complete animal.
Independent variable is the size of fragment (1/2, 1/4, …)
Dependent variable is time to fully regenerate.
Controlled variables are all environmental variables including temperature, pH, light.
Constants are experiment method, procedures and growth medium.
* * *
For the second question pH is the independent variable. (fragment size will also be added to the list of constants)
For the third question light is the independent variable. (fragment size will also be added to the list of constants)
For the fourth question temperature is the independent variable. (fragment size will also be added to the list of constants)
Hypothesis:
Based on your gathered information, make an educated guess about what types of things affect the system you are working with. Identifying variables is necessary before you can make a hypothesis.
Following are sample hypothesis for the four questions that I have proposed:
- The larger a fragment of planarian, the less it takes to regenerate.
- pH has no affect on the rate of regeneration of planarians.
- Planarians regenerate faster in darkness.
- Planarians regenerate faster in warm water.
Note that the results of your experiments may show that your hypothesis has been wrong. Also it is a good practice to offer an explanation with each hypothesis describing why you are adopting this hypothesis.
Experiment Design:
Design an experiment to test each hypothesis. Make a step-by-step list of what you will do to answer each question. This list is called an experimental procedure. For an experiment to give answers you can trust, it must have a “control.” A control is an additional experimental trial or run. It is a separate experiment, done exactly like the others. The only difference is that no experimental variables are changed. A control is a neutral “reference point” for comparison that allows you to see what changing a variable does by comparing it to not changing anything. Dependable controls are sometimes very hard to develop. They can be the hardest part of a project. Without a control you cannot be sure that changing the variable causes your observations. A series of experiments that includes a control is called a “controlled experiment.”
Experiment 1: Planaria Regeneration Rate
Introduction
Many animals are capable of regeneration of lost parts to some extent. In this experiment we will examine regeneration in Dugesia sp. (a planarian), a common freshwater flatworm. These animals have considerable powers of regeneration, and their ability to regenerate has been investigated in great detail. The body is polarized on the anterior-posterior axis. In other words, if you cut off the head the head will grow a new body, and the body will grow a new head. Also, anterior regions grow fastest.
Materials
- 2 complete petri dishes (top and bottom)
- Dugesia (from pond or Carolina L210)
- razor blades
- droppers
Preparation
You must have fully grown planaria for this experiment. If you are growing planaria from culture, you must follow the instructions that you receive with the culture to grow planaria so they will be ready for this experiment. If you are collecting planaria from a pond, you must isolate planaria from algae and other invertebrates that may exist in water.
How to isolate planaria from pond water?
Add tap water to the bottoms of a petri dishes. Take some pond algae and put it in the bottom of another petri-dish. Use an eye dropper to suck up a planarian and put it in the first petri dish. Be sure not to include other visible invertebrates as they may be predators of Dugesia. Collect 20 individuals if possible (keep track by counting them!). Throw away the algae, wash the plate and eye dropper. keep your 20 isolated planarians for this experiment.
Experiment:
Half fill 2 Petri dishes with simulated pond water or aged tap water. This water contains most of the nutrients but none of the micro-organisms of ordinary pond water.
Use an eye dropper to transfer 10 Planaria to each petri-dish.
Label one petri-dish containing 10 planaria and somewater as “control” and close the top.
Place the other petri-dish on ice so planaria will become motionless.
With the scalpel or razor blade, quickly cut each Planaria in half. Cuts can be transverse, or longitudinal. Cut exactly half of the worms. Leave both halves in the petri-dish.
Close the lid and label this petri-dish with “Cut“.
Place the Petri dishes in a dark place at room temperature or in an incubator.
Examine the Planaria for a few minutes every 3 days. Remove any dead sections immediately. Examine the Planaria under a magnifier or dissecting microscope and make sketches to show the changes. Note how many have full, partial, or no regeneration.
Record your observations in a table like this:
Number of live worms in petri-dishes
| Date | No regeneration | Partial Regeneration | Full regeneration | Control |
References
Any invertebrate zoology text.
Rose, S.M. 1970. Regeneration. Appleton-Century-Crofts, NY
Moraczewski, J. 1977. Asexual reproduction and regeneration of Catenula. Zoomorphologie 88:65-80.
Whitten, R.H. and W.R. Pendergrass. 1980. Carolina protozoa and invertebrates manual. Carolina Biological Supply Co., Burlington, NC. 34 pp.
Experiment 2: Effect of pH on the rate of Regeneration in planaria
Introduction:
Scientists often want to know the affect of environmental factors on the rate of reproduction in different animals. Regeneration for planaria is also a way of asexual reproduction, so it well qualifies for such studies.
Procedure:
This experiment is identical to the previous experiment. Instead of 20 planaria, use 40 planaria in 4 petri-dishes (10 planaria per dish). One dish will be control and 3 others will have cut planaria. These last 3 dishes will have three different pH. One will be normal water pH. The other will be acidic (by one drop of acetic acid) and the last one will be alkaline (by one drop of ammonia solution).
Record 3 results table, one for each pH (Alkaline, normal, acidic). Use the same control for all three results tables.
More Experiments: Effect of other factors such as temperature or light on the rate of Regeneration in planaria or sponges
Many other experiments can be designed to test the effects of certain factors (independent variables) on the rate of regeneration on planarians and sponges.
The structure and procedures of most such experiments are similar to the experiment 1 and 2 described above.
Sponge growth Experiment
Introduction:
Sponges are very simple animals whose cells are not organized into discrete tissues. They are capable of incredible regeneration. The classic experiment to demonstrate this property is to force a sponge through a fine mesh to separate the cells. After a while, the cells re-aggregate by type and reform the sponge’s body.
Procedure:
Obtain a piece of live Microciona or any other marine sponge. Press it through a metal or nylon mesh into seawater. This will break the sponge in small clumps of cells. These clumps will remain suspended in sea water. Examine a drop of the suspension under a compound microscope or magnifier glass. You will see small clumps of cells.
Prepare a series of dilutions in seawater from the original suspension so that you have samples of decreasing densities. Make your dilutions 100%, 50%, 25%, and 10% solutions of the original suspension.
Get 8 identical beakers or glass cups and half fill two beakers with each of the dilutions. Label the beakers with the dilution. Also label one beaker of each dilution as control. (So you will have one control for each dilution).
Cover the control beakers and place them in refrigerator. Keep other beakers at room temperature (72-80ºF) and use an aquarium pump to delver air to these cups a few hours each day.
Make observations of cell clumps every 2 days using a microscope or magnifier glass. If the pieces are still very small, you may use a pipette to transfer a drop of the suspension to a microscope slide and then view it under a the microscope.
Does temperature affect regeneration of sponges?
Materials and Equipment:
- 2 complete petri dishes (top and bottom)
- Dugesia (from pond or Carolina L210)
- razor blades
- droppers
Results of Experiment (Observation):
Experiments are often done in series. A series of experiments can be done by changing one variable a different amount each time. A series of experiments is made up of separate experimental “runs.” During each run you make a measurement of how much the variable affected the system under study. For each run, a different amount of change in the variable is used. This produces a different amount of response in the system. You measure this response, or record data, in a table for this purpose. This is considered “raw data” since it has not been processed or interpreted yet. When raw data gets processed mathematically, for example, it becomes results.
Calculations:
No calculation is required for this experiment.
Summary of Results:
Summarize what happened. This can be in the form of a table of processed numerical data, or graphs. It could also be a written statement of what occurred during experiments.
It is from calculations using recorded data that tables and graphs are made. Studying tables and graphs, we can see trends that tell us how different variables cause our observations. Based on these trends, we can draw conclusions about the system under study. These conclusions help us confirm or deny our original hypothesis. Often, mathematical equations can be made from graphs. These equations allow us to predict how a change will affect the system without the need to do additional experiments. Advanced levels of experimental science rely heavily on graphical and mathematical analysis of data. At this level, science becomes even more interesting and powerful.
Conclusion:
Using the trends in your experimental data and your experimental observations, try to answer your original questions. Is your hypothesis correct? Now is the time to pull together what happened, and assess the experiments you did.
Related Questions & Answers:
What you have learned may allow you to answer other questions. Many questions are related. Several new questions may have occurred to you while doing experiments. You may now be able to understand or verify things that you discovered when gathering information for the project. Questions lead to more questions, which lead to additional hypothesis that need to be tested
Possible Errors:
If you did not observe anything different than what happened with your control, the variable you changed may not affect the system you are investigating. If you did not observe a consistent, reproducible trend in your series of experimental runs there may be experimental errors affecting your results. The first thing to check is how you are making your measurements. Is the measurement method questionable or unreliable? Maybe you are reading a scale incorrectly, or maybe the measuring instrument is working erratically.
If you determine that experimental errors are influencing your results, carefully rethink the design of your experiments. Review each step of the procedure to find sources of potential errors. If possible, have a scientist review the procedure with you. Sometimes the designer of an experiment can miss the obvious.
References:
List of References
