PRACTICAL MANUAL: Plant Growth Regulators Gibberellic Acid-3 (GA-3)

Introduction

Gibberellic Acid-3 (GA-3) is a naturally occurring plant growth regulator that causes a variety of physiological effects, including the stimulation of seed germination and stem elongation. GA-3 occurs naturally in the seeds of many species and is produced commercially by growing Gibberella fujikuroi fungus cultures, followed by extraction and purification.

Presoaking seeds in a GA-3 solution can trigger rapid germination in many types of highly dormant seeds. In this experiment, we will observe how GA-3 affects the growth of two different types of seeds:

• Endospermous seeds (Corn, Zea mays): Seeds that store nutrients in the endosperm.

• Non-endospermous seeds (Long bean, Vigna unguiculata subsp. sesquipedalis): Seeds that store nutrients in the cotyledons.

Objectives

1. To study and compare the influence of GA-3 on the seedling growth of endospermous (corn) and non-endospermous (long bean) seeds.

2. To gain practical experience in setting up germination assays and maintaining a sterile at-home growing environment.

3. To develop skills in experimental data collection, statistical analysis, and scientific data visualization through class collaboration.

Materials and Methods

This is a collaborative class experiment. There are 6 treatment combinations in total based on the 2 seed types and 3 GA-3 concentrations (0 ppm, 200 ppm, and 400 ppm).

Your group will choose ONE of the following 6 treatment combinations to be responsible for:

1. Corn + 0 ppm (Control)

2. Corn + 200 ppm

3. Corn + 400 ppm

4. Long bean + 0 ppm (Control)

5. Long bean + 200 ppm

6. Long bean + 400 ppm

Procedure:

1. Seed Viability Test: Place a handful of your assigned seed type in a beaker of water. Discard any seeds that float to the top (these are dead or empty). Select 10 to 12 sinking, viable seeds for your experiment.

2. GA-3 Treatment: Soak your selected 10-12 seeds in your assigned GA-3 solution for exactly 1 hour.

3. Petri Dish Setup: Take 1 clean petri dish and line the bottom with a paper towel. Ensure your group name, seed type, and assigned GA-3 concentration are clearly written on the lid.

4. Plating: After the 1-hour soak, transfer the treated seeds into your prepared petri dish.

5. Moistening: Wet the paper towel using filtered water. Ensure the tissue is thoroughly moistened but the seeds are not entirely submerged. Cover the dish with its lid.

6. At-Home Maintenance: Bring the petri dish home and place it near a window with indirect sunshine. Check it regularly. Whenever the paper towel begins to dry, remoisten it strictly with filtered water to prevent fungal or bacterial contamination.

7. Data Collection: Once germination begins, measure and record the seedling shoot length in centimeters daily for a duration of 8 days.

Results

Record your group's daily measurements and calculate the average shoot length of your 10-12 seeds. At the end of the 8 days, you will share your group's averages with the rest of the class so everyone can fill out the complete tables below.

Table 1: Average Growth of Endospermous Seed (Corn Zea mays)

Days After Planting	Control (0 ppm)	Medium (200 ppm)	High (400 ppm)
0
1
2
3
4
5
6
7
8

Table 2: Average Growth of Non-endospermous Seed (Long bean, Vigna unguiculata subsp. sesquipedalis)

Days After Planting	Control (0 ppm)	Medium (200 ppm)	High (400 ppm)
0
1
2
3
4
5
6
7
8

Data Analysis & Discussion

1. Class Data Compilation & Statistical Visualization: Compile your final day (Day 8) data with the rest of the groups to compare the final growth outcomes between all 6 treatment combinations.

   Using software like Excel, Google Sheets, or GraphPad Prism, construct a grouped bar graph to visualize the class results. Your graph must meet standard scientific formatting requirements, including:

   • Properly labeled axes: The Y-axis must include the unit of measurement (e.g., "Final Shoot Length (cm)"), and the X-axis should clearly denote the GA-3 treatments.

   • Clear legends: Include a legend to easily distinguish between the endospermous (corn) and non-endospermous (long bean) data series.

   • Error bars: Calculate and include error bars representing the Standard Error (SE) of the mean for each treatment group to show data variability.

   • Figure caption: Below the graph, provide a brief caption that states the sample size (n-number) used to calculate the averages and standard error (e.g., "n = 10-12 seeds per treatment").

2. Discussion:

   • Based on the completed bar graph, discuss the physiological responses that occurred from the different GA-3 treatments. Did increasing the concentration yield a linear increase in growth?

   • Conclude whether the endospermous (corn) or non-endospermous (long bean) seeds were more responsive to the exogenous GA-3. Discuss why this might be the case, referring to the structural and physiological differences between how these two seed types mobilize their stored food reserves during germination.