Engaging inexpensive hands-on activities using Chlamydomonas reinhardtii (a green micro-alga) beads to teach the interplay of photosynthesis and cellular respiration to K4-K16 Biology students

Abstract

Background: Photosynthesis and cellular respiration play major roles in energy metabolism and are important Life Science topics for K16 Biology students. Algae beads are used for photosynthesis and cellular respiration labs. Currently there are a few companies that sell biology educational kits for making algae beads using non-motile green micro-algae to introduce students to photosynthesis. These kits are expensive and, do not come with detailed guidelines for trouble shooting and customizations for different grade levels. Chlamydomonas reinhardtii is a motile green micro-alga and is an excellent model system for photosynthesis studies. In this article, we are presenting the work conducted in the student-driven, American Society of Plant Biologists-funded, Plant-BLOOME educational outreach project. This project is a supervised collaborative effort of three undergraduates and one high school student. We have generated a protocol which can be used to make Chlamydomonas beads. We have used these beads to design two simple and inexpensive plant biology hands-on activities. These laboratory activities have been customized to teach the interplay of photosynthesis and cellular respiration to K4-K16 Biology students.

Methods: Chlamydomonas beads were used for two different laboratory activities that involved monitoring pH changes over time using a pH indicator. Our first activity centers on making and, using light-powered algae bead bracelets to monitor dramatic color/pH changes over time when exposed to darkness or light. Our second activity employs strain-specific algae beads with approximately equal cell numbers to conduct comparative photosynthesis and cellular respiration studies in two Chlamydomonas strains namely, wild type, 4A+ and, a high light-sensitive, photosynthetic mutant, 10E35/lsr1a.

Results: We optimized our experimental protocol using algae beads in a 5.5 mL screw capped glass vials before performing the same experiment in algae bead bracelets. We found that the algal cell density/bead, water type used in the experiment and, the duration of dark exposure of algal beads can affect successful implementation of the lab activities. Light-powered algae bead bracelets showed dramatic color/pH changes within 3 h upon exposure to light or darkness. These bracelets could be switched back and forth between darkness and light multiple times within 48-72 h to display color/pH changes, provided prior dark exposure time did not exceed 9 h. Our comparative studies of photosynthesis and cellular respiration in 10E35 and in 4A+ showed that relative respiration rate and photosynthetic rate is higher and lower in 10E35, respectively, compared to that in 4A+. Additionally, 10E35 failed to display the expected photosynthesis-induced pH/color changes in the light after prolonged exposure to darkness which indicated that prolonged dark exposure of 10E35, hindered photosynthesis.

Keywords: Algae bead bracelets; Algal beads; Biology education; Cellualr respiration; Chlamydomonas; Green micro-alga; Light-powered algae bracelets; Photosynthesis; Plant biology.

Figure 1. Photosynthesis and cellular respiration-induced pH/color changes in vials containing Chlamydomonas 4A+ strain beads in de-ionized water and tap water.

C stands for control vials which do not contain algae beads and E stands for experimental vials containing algae beads. (A) Color changes in de-ionized water. (B) Color changes in tap water. Vials were exposed to light and darkness for 2 hours. Algal beads had approximately 2 × 10⁶ cells/beads and 8 algal beads were used per experimental vial. All statistical analyses can be found in https://doi.org/10.6084/m9.figshare.12344024.v1, Data S1 and S2 and Table S1.

Figure 2. Effect of cell numbers per Chlamydomonas 4A+ strain bead on photosynthesis and cellular respiration-induced pH/color changes in tap water.

“C” stands for control vials that do not contain algae beads. “E” stands for experimental vials containing algae beads. (A) Color changes in experimental vials that contained beads which had approximately 4 × 10⁶ cells/bead. (B) Color changes in experimental vials that contained beads which had approximately 2 ×10⁶ cells/bead. Light and dark exposure of vials was for 2 hours. Eight algal beads were used per experimental vial. All statistical analyses can be found in https://doi.org/10.6084/m9.figshare.12344024.v1, Data S1 and S2 and Table S2.

Figure 3. Indirect monitoring of carbon dioxide percentage inside the 4A+ bead bracelet under constant light and darkness using bicarbonate indicator.

(A) Control algal bracelet that has not been exposed to light or to darkness (zero time point). (B) An algal bracelet that was exposed to darkness for 3 hours. (C) An algal bracelet that was exposed to light for 3 hours. Water color change was monitored in dark and light. Algal beads had approximately 2 × 10⁶ cells/bead. Number of algal beads in the control, dark- and light-exposed bracelets were 32, 38 and 38, respectively. All statistical analyses can be found in https://doi.org/10.6084/m9.figshare.12344024.v1, Data S1 and S2 and Table S3.

Figure 4. Time course monitoring of photosynthesis-induced color/pH changes in the dark-adapted 4A+ bead bracelet when shifted to light.

(A) An algal bracelet that was dark adapted for 4 hours. (B) Dark-adapted bracelet exposed to light for 1 hour. (C) Dark-adapted bracelet exposed to light for 2 hours. (D) Dark-adapted bracelet exposed to light for 3 hours. (E) Dark-adapted bracelet exposed to light for 4 hours. Algal beads have approximately 2 × 10⁶ cells/bead. Number of algal beads in the bracelet was 15. Bicarbonate indicator was used as a pH indicator.

Figure 5. Time course monitoring of cellular respiration-induced color/pH changes in the light-adapted 4A+ bead bracelet when shifted to darkness.

(A) An algal bracelet that was light adapted for 4 hours. (B) Light-adapted bracelet exposed to dark for 1 hour. (C) Light-adapted bracelet exposed to dark for 2 hours. (D) Light-adapted bracelet exposed to dark for 3 hours. Algal beads had approximately 2 × 10⁶ cells/bead. The bracelet contained thirty-six 4A+ strain beads. Bicarbonate indicator was used as a pH indicator.

Figure 6. Effect of prior dark exposure duration on photosynthesis-induced color/pH changes in 4A+ bead bracelet in light.

(A) An algal bracelet that was dark adapted for 9 hours. (B) 9 hours-dark adapted-bracelet shifted to light for 4 hours. (C) An algal bracelet that was dark adapted for 15 hours. (D) 15 hours-dark adapted-bracelet shifted to light for 12 hours. Algal beads had approximately 2 ×10⁶ cells/bead. Both bracelets contained 38 beads. All statistical analyses can be found in https://doi.org/10.6084/m9.figshare.12344024.v1, Data S1 and S2 and Table S4.

Figure 7. Comparative studies of photosynthesis and cellular respiration-induced pH/color changes in wild type 4A+ and 10E35 bead vials under light and darkness.

(A) Control, 10E35 and 4A+ bead vials before light exposure. (B) Control, 10E35 and 4A+ bead vials after 30 minutes of light exposure. (C) Control, 10E35 and 4A+ bead vials after 1 hour of light exposure. (D) Control, 10E35 and 4A+ bead vials before dark exposure. (E) Control, 10E35 and 4A+ bead vials after 30 minutes of dark exposure. (F) Control, 10E35 and 4A+ bead vials after 1 hour of dark exposure. Algal beads of each strain had approximately 2 × 10⁶ cells/bead. Eight beads of each strain were used per experimental vial for the experiment. All statistical analyses can be found in https://doi.org/10.6084/m9.figshare.12344024.v1, Data S1 and S2 and Table S5.

Figure 8. Time course monitoring of cellular respiration-induced color/pH changes in the dark in 4A+ and 10E35 bead vials that were adapted to light for 4 hours.

These vials are the light-adapted vials from the Fig. 7 experiments. (A) Light-adapted control, 10E35 and 4A+ bead vials before dark exposure. (B) Control, 10E35 and 4A+ bead vials after 15 minutes of dark exposure. (C) Control, 10E35 and 4A+ bead vials after 30 minutes of dark exposure. (D) Control, 10E35 and 4A+ bead vials after 45 minutes of dark exposure. (E) Control, 10E35 and 4A+ bead vials after 1 hour of dark exposure. All statistical analyses can be found in https://doi.org/10.6084/m9.figshare.12344024.v1, Data S1 and S2 and Table S6.

Figure 9. Time course monitoring of photosynthesis-induced color/pH changes in the light in 4A+ and 10E35 bead vials that were adapted to darkness for 6 hours.

These vials are the dark-adapted vials from the Fig. 8 experiments. (A) Dark-adapted control, 10E35 and 4A+ bead vials before light exposure. (B) Control, 10E35 and 4A+ bead vials after 30 minutes of light exposure. (C) Control, 10E35 and 4A+ bead vials after 1 hour of light exposure. (D) Control, 10E35 and 4A+ bead vials after 2 hours of light exposure. (E) Control, 10E35 and 4A+ bead vials after 3 hours of light exposure. (F) Control, 10E35 and 4A+ bead vials after 48 hours of light exposure. All statistical analyses can be found in https://doi.org/10.6084/m9.figshare.12344024.v1, Data S1 and S2 and Table S7.