Main Page/Culturing Chara

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Culturing Chara

Chara lifeform 01.png

Overview.

Chara is used for teaching labs in Biology and Biophysics. In BIOL2010 (Plant Biology), it is used for diversity labs. In BIOL2070 (Methods in Cell and Molecular Biology), it is used to explore molecular motors. In BPHS4090 (Biophysics II), it is used to measure action potentials [[1]]. Here, we provide a description of how to culture Chara.
Raising Chara.png

Chara australis planting technique.

Chara is a multicellular freshwater algae which roots itself in soil, and grows upward to the surface of the water. So, our aquarium tank cultures of Chara normally uses a soil base covered with a growth medium. The size of the tanks will affect the quantity of the algae. We use either small (20 cm width by 41 cm length by 26 cm depth) or large (27 cm width by 49 cm length by 30 cm depth) aquaria tanks.


Soil preparation.

We obtain our soil from a local woodlot (Boyer Woodlot —northwest corner at the edge of a path leading to the road, click on the map).
Chara soil location.png
The soil surface is scraped bare of vegetation (leaves etc.) and collected to a depth of about 2 cm. The soil is mostly silt with some clay and organic material. We screen it through 1 cm mesh to break clumps, then sterilize it by autoclaving. The soil is placed in a metal container, covered with aluminum foil, and autoclaved for 90 minutes using a liquid cycle (slow exhaust). When the cycle is complete, the soil is removed from the autoclave and left overnight at room temperature to cool. The next day it is re-autoclaved (90 minutes, liquid cycle) and cooled to room temperature. If the soil is stored for an extended period, it is advised that the soil be allowed to dry out, to minimize the growth of fungal molds.


Planting Chara.

The soil is added to the aquarium tanks to a depth of about 2–3 cm, and saturated with Broyer-Barr medium (see stock make-up, below). Sections of Chara are cut from an existing culture. The explants include the plant top and 2 to 4 internodes (total length about 10–20 cm). The Chara explants are buried in soil by making a groove in the soil by hand and gently placing the explant in the groove (horizontally), then covered with additional ‘sprinkle’ of soil. The apex is allowed to extend above the soil surface. Broyer-Barr medium is added gently by pouring slowly onto cheese cloth adhering to the aquarium tank wall to minimize any disturbance of the soil. We usually start with a liquid depth of about 3 cm. Over time, as the Chara grows, the height of the medium is increased (usually every week or so), eventually to the top of the tank. An alternative planting technique is to cut the plant top with 1 mature internode. A whorl cell at the bottom of the plant top is held with forceps, and the bottom whorl inserted into the soil, leaving the top in solution. Continued growth of the plant apex is often observed within days (see montage below). Sometimes, above-ground plant parts will die off (turning black). This is normal: new sprouts will emerge from the underground Chara plantlets. Once planted, the tanks are covered with plexiglas covers, or plastic wrap (to minimize evaporation).

Chara Growth montage.png

Growth of Chara The montage is snapshots from a time lapse movie (every five hours). For scale, the internode cell diameter is about 1.5 mm. Movies in quicktime format are available at http://www.yorku.ca/planters/movies.html#Chara.


Snail 00.png
Snails can be transplanted into the tanks at the same time or soon thereafter. The snails will feed on other algae that pioneer the tank, and will climb along the Chara internodes, feeding on epiphytic algae that attach to the internodes.


Chara tanks.png
Long-term, the solution can be replaced (to minimize turbidity caused by planktonic micro-algae). This is done by siphoning from the bottom of the tank using Tygon tubing, while replacing the media at the surface on the opposite side of the tank (using cheesecloth to minimize disturbance of the soil). It is even possible to use RO (reverse osmosis de-ionized) H2O since the soil provides the salts and micro-nutrients required for growth of the autotrophic Chara. Avoid using tap water (due to chlorine). Flushing the tanks is usually done fairly regularly with new cultures (about once a week), but stopped when the Chara is well established (it usually takes 2-3 months to obtain dense growth of the Chara).

Illumination should not be too bright (to discourage growth of planktonic algae and cyanobacteria). Normally, photosynthetically-active radiation (PAR) should be about 20 µmole m–2 s–1, measured with a radiometer (less intensity is insufficient for growth, greater intensity encourages growth of cyanobacteria). We normally use T8 fluorescent lamps with the highest color temperature we can find (4100 K for 24 inch or 6500 K for 48 inch lamps) on a 12 hour dark / 12 hour light cycle. We have noticed that the lamps become dimmer over time, and now replace the bulbs every 1-2 years to ensure sufficient light intensity.

Media Make-up

Below is a make-up list for Broyer-Barr media, available as a png image file.

Media makeup.png

Chara Disposal.

In keeping with the view that organisms should not be introduced into the environment, we routinely destroy Chara cultures when the cultures are no longer required. To do this, we siphon the medium from the tank, add household bleach (5% sodium hypochlorite) so that the soil is complete saturated and then cover the tank top. After 1 day, the Chara will have died, as evidenced by its straw-like color due to chemical bleaching of the chlorophyll. At this point we discard in the greenhouse bins for soil disposal, flush the tank with water, and clean with a standard glassware detergent to remove algal biofilms from the glass sides of the tank. Now, all is ready for the next round of Culturing Chara!

We thank Professor Mary Bisson (University at Buffalo) for getting us started with Chara in support of teaching exercises.

  • Created and maintained by RR Lew, Biology Department at York University
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