Useful Methods 2: Sterilization of Duckweed

Put the plants in a plastic tube (Falcon), depending on the size of the plants having a volume of 15mL or 50mL.

Dilute the commercial available “Eau de Javel” in water for different concentrations and treat the fronds for different periods by gentle shaking:

Wolffia, Wolffiella : 1 – 3%, 2, 3, 4, and 5 min.

Lemna : 3 – 5%, 3, 4, 5, and 6 min.

Landoltia, Spirodela : 5 – 10%, 3, 5, 7, and 9 min.

Sterilize Landoltia fronds in a 0.24% solution of NaOCl using “Eau de Javel".

Transfer the plants into Erlenmeyer flasks with nutrient medium containing sugar (50millimolar (mM) glucose or 25millimolar (mM) sucrose) (e.g. 180mL per flask) without further washing in sterile water.

After a few days, better after 14 days, the medium remains clear when all bacteria and fungi are killed.

Wash the plants in 70% ethanol before bathing them in “Eau de Javel”. However this does not improve the rate of success.

Treat the 4 Erlenmeyer flasks with fronds at different times in the bleaching bath.

In the cultivation room at 25°C follow the development of the plants in the Erlenmeyer flasks.

Transfer the sterilized plants on Agar in Petri dishes.

We also use other commercially available preparations like “Danklorix” (CP GABA, Hamburg, Germany), available at least in Austria and Germany. This product contains 2.8% NaOCl and can be used in the same way as “Eau de Javel”.

Limited experience exists with commercial available sodium hypochlorit (NaOCl) solution. However, the concentration is much higher (typically 12%) and dilute it accordingly.

Moreover, as it is free of surfactants, it is useful to add a small amount of a mild detergent like 0.5% of Triton X-100 to get the surface of the whole plant body wetted.

There are other beaching agents like “Lizol” (Reckitt Benckisier, Parwanno, Himachel Pradesh, India) having e.g. 0.6% NaOCl. However in this case, a very high concentration of surfactants is included. We were not able to wash the surfactants properly away from the surface of the plants and the recovering rates were very low.