Hormone-Induced Plant Propagation Lab Using Carrot Root Cultures

C. Anthony Finney

Flushing International High School, Queens

Summer Research Program for Science Teachers

August 2008

Grade Level: 9th and 10th Grades  

Unit: Lab activity at end of plant biology unit, with ties to hormones, reproduction, development ecology and human impact themes

Objectives:

After completing this lab, students will develop:

SCIENCE SKILLS, including:

Following good lab practice (safety, aseptic technique, etc.)

Identifying components of an experiment (hypothesis, control, etc.)

Designing an experiment to test a hypothesis (see Introduction note 3)

SCIENCE CONTENT UNDERSTANDING, including:

The effect of hormones on tissue development

Basic cell theory, including the cellular basis of life, and the cellular origin of new cells.

Compare the benefits and disadvantages of genetically identical crops 

Materials:

Fresh unprocessed carrots

Cutting tools: scalpels, etc.

Fume hood

70% EtOH

20% Bleach solution

Clock or timers

Beaker or other liquid waste receptacle

Vegetable peeler

Synthetic auxin (plant hormone) such as Rootone^Ò (http://www.gardentech.com/)

Growth media - e.g. Agar

Transfer loops, orangewood sticks or sterile swabs

Hand lenses or dissecting microscope

Petri dishes

Flame source (to sterilize instruments frequently)

Liquid detergent

Parafilm (or tape)

Introduction:

Propagation is a centuries-old practice by which desirable plant specimens are asexually reproduced to produce more copies of that organism for human use.  Correctly dividing each tissue to yield two viable organisms from a single parent is a skill any good horticulturist has mastered.  In the 1920s and 30s, however, scientists extracted and described a hormone which help plants to grow in the root-down way we are familiar with.  The hormone, termed auxin, is critical in root development.  This lab uses a commercial synthetic preparation of that hormone to stimulate development in a prepared plant sample.

To Note:

1. This lab is often used in an undergraduate botany class.  Because it uses EtOH, flaming of sharp instruments and aseptic procedure, well-developed lab safety skills should be instructed

2. Although the procedure itself requires only an hour, the development of new plant tissues from the sections prepared require time to grow.  Therefore, this lab is intended as a long-term, intermittent project suited to linking several units of study, and developing student skill at making observations over time.

3. To include an investigatory element to the lab procedure described below, other variables can be introduced (caffeine, sugar, etc.) and class results compared at the conclusion of the lab.

4. Extensions envisioned for bridging this lab to other units include:

              Biotechnology / Biomedical Ethics

Procedure:

1.      Select a fresh carrot, free from cracks or other damage from those supplied.  In your lab notebook, make qualitative and qualitative observations of your specimen.  Use tools, including hand lenses and measuring tape/rulers to add detail to your observations.

2.      Rinse your carrot in the sink to remove any loose dirt.

3.      Cut off the top and bottom on the carrot and peel off the skin with a peeler.

4.      Trim the peeled carrot into 10cm sections and place them into a beaker

5.       In the flow hood, cover the carrot sections with 70% EtOH and sitr for 5 minutes.

6.       Pour off the EtOH into a waste beaker.

7.      Immediately cover the carrot sections with a 20% bleach solution + a drop of detergent for approximately 20 min, then decant off the bleach solution.

8.      Rinse the carrots three times with sterile water, covering the sections and stirring for 1-2 min each time.

9.      Transfer a section into a sterile dish.  Using scalpel and forceps, cut small, equal- sized pieces of root, making sure to include the cambium (the innermost ‘ring’ of the root).  Flame Instruments often.  You will need 8 total pieces.

10.  Using the forceps, place 4 of the carrot pieces on the provided agar media plate

11.  Label the plate ‘Control’ with your name and the date

12.  Using the forceps, touch 1 or two sides of each of the last four carrot pieces in the powdered plant hormone to coat it.

13.  Place these final 4 carrot pieces on the provided agar media plate

14.  Label the second plate ‘Hormone’ with your name and the date

15.  Seal each dish with parafilm, and incubate in the dark.

16.  Each week, record your observations on the carrots’ development in your lab notebook

17.  After 4 weeks, subculture the callus onto the same medium, discarding any dead tissue.

Data Collection:

Data collection includes qualitative / quantitative data on starting carrot tissue cultures, and follow-up observation of plant cultures after each week’s growth

Analysis Questions:

After DAY1

1.      What does it mean to use ‘aseptic’ procedure?  Why do you think it is important to use aseptic procedure in this lab?

2.      What are some similarities between how humans and plants are able to reproduce?  What are some differences?

3.      What is a hormone, and why is it important in living things?

4.      Describe the differences you expect to see on your plates in a week.  What factors do you think will be most important in causing these results you predict.

After DAY 5 (a week later)

5.      What differences do you observe between each of the carrot pieces today and what they looked like a week ago?  What changes can you see in each of the samples?  Refer to your lab notebook if you need help remembering the original appearance.

6.      What differences can you find between the ‘Control’ and ‘Hormone’ plates?  How can you explain this result?

7.      Describe anything OTHER than carrots you see growing on the plates you prepared.  What is it, and how do you think it got there?

8.      Refer to your lab notebook response to question 3.  Was your prediction correct or incorrect? 

9.      This lab involved the use of plant hormones.  Select a human hormone to do further research on.  In your response, name the hormone, tell what tissues produce it, and describe how it affects the body

10.  How could this activity be used for practical purposes?  What are the disadvantages and benefits of using this procedure to grow new plants?

11.  How is it possible to make an entire new organism from one small piece of that organism?

Standards:

This lesson plan addresses the following local, state and national standards: