Germination in vitro:
future for the
Victoria amazonica seeds?
Royal Botanic Gardens, Kew
by Renata Mazzini
Click images to enlarge
Acknowledgements - References - About the author
Victoria amazonica (Nymphaeaceae), native to the Amazon forest of South America, was first brought to England in the 1800's. It was named after Queen Victoria, Victoria regia, and it is impressive for the size of its leaves and flowers. There are two species of the genus Victoria - amazonica and cruziana, and several hybrids, the most famous being the 'Longwood Hybrid', created back in the 1960's, at the Longwood Gardens, in USA.
Some growers keep the plant for several years, but it is still considered an annual in the Northern Hemisphere. In its natural habitat, it can survive for many years, probably because of the warm and humid weather all year long. It is only reproduced from seed, and that is why it is important to sow and have seeds available every year. It is a great attraction at Kew Gardens, and lots of people come to the gardens only to see this natural wonder.
Victoria amazonica is the only species studied in this project, because it requires some more specific conditions for germinating than the other ones: it needs a warmer temperature, around 30°C. Victoria cruziana grows at cooler ambient, and the hybrids are much hardier. The amazonica species is always the slowest and most difficult to germinate.
At Kew, the seeds are nicked (see "Breaking the dormancy") before sown in loam, and kept in a tank at 30°C, in a zone where the minimum air temperature is 18°C, and the maximum 22°C. Because it takes from 3 to 4 moths to get the plants ready for their final position, the seeds have to be sown in February, to have them ready for the summer display. Unfortunately, the day length in February is still shorter than that considered ideal, and so it can take longer to germinate.
For this project, some experiments were done using two germination
in vitro components: one with Gibberellic Acid (GA3), and another one with Agar. The objective
is to test if these materials can improve the germination rate,
and ensure a good number of plants.
The factors that can influence in the Victoria amazonica seed germination process are: light, germination and storage temperature, interruption of dormancy, compost used and seed age.
Many tests related to its germination process have already been done in botanical and private gardens, but unfortunately, there is no agreement between them.
§ The experiment:
The seeds came in three different lots from Kit and Ben Knotts, the Paradise Garden's owners, a private garden in USA. There were 40 seeds from each.
Among all of the seeds, 48 (16 from each lot) were sown in loam, 12 in water (4 from each lot), 24 in Gibberellic Acid (8 from each lot) and 24 in agar (8 from each lot). Half of the seeds were nicked (see "Breaking the dormancy"), to compare if this method really improves the germination rate. They were checked every day during 35 days. This is the maximum that it should take for a Victoria seed to germinate. In some places in USA, as at the Paradise Garden or at the New York Botanical Garden, nicked seeds have started to germinate in 4 and 2 days, respectively. At Kew, they usually start around 10 days.
Seeds sown in loam, inside the plastic bags and in small flasks
with Gibberellic Acid (GA3) and Agar
§ The temperature:
At the Paradise Garden and people from The Victoria Conservancy, both in USA, store their seeds at 24°C, while other ones, like at the National Botanic Garden of Belgium and also at Kew, store theirs at 13°C and 14°C, respectively. The Knotts also say that "any chilling of amazonica seeds GREATLY reduces viability." The seeds have to be stored in water, in the dark, otherwise some premature sprouting can occur. The seed germination temperature is the same in everywhere, from 28°C to 32°C.
The seed storage temperature was not tested. They should have been at the same temperature since they were collected, although they did suffer some cooling when they arrived. The seeds were stored at 24°C, but when they arrived, all of them were in a fridge at 14°C during 24 hours, and then half of them were moved to a room for 2 days, where the average temperature was 18°C, and then were sown in loam. Half of the seeds were in the same zone as the sown ones, inside the glasshouse, where they were sown in GA3 and Agar 13 days later. The water temperature in the tank, for the seeds to germinate, was 30°C. It does not seem that this temperature variation influenced in the results, because the germination rate was very variable.
There is a method that improves the Victoria seeds germination rate. It is called "seed nicking". The operculum (the "door" which the embryo germinates through) is lifted off with a sharp knife, being very careful not to damage the embryo, which is just beneath it. This is a very difficult process because it requires much attention and specialisation, but it improves the germination rate, making it faster, because the embryo does not have to break the hard seed coat to germinate. Care must be taken not to let the seeds dry out.
Some other growers just scarify the seeds, or leave the seeds in a weak bleach solution, but at the Longwood Gardens, USA, this last method does not seem to improve the germination rate. It might make the seedlings more resistant to pests and diseases. The Knotts did not see any difference either.
§ The compost used:
After testing different composts, the Knotts found that the best "compost" to germinate the seeds is only water, in small plastic bags, but they recommend that 25% of the water has to be changed twice a week. During these 35 days, the water was changed only once, and it appears that germination was still improved. For storing and germination, it can be tap water, but after that, it should be treated water, either because the seedlings need some nutrients to keep growing, or because there are some components in tap water, such as chlorine and copper, which are toxic to them. At Kew, composts like pure sand, and then sand + loam were tested in the past. Nowadays, the seeds are sown in soaked loam, but for this project, seeds were sown in both loam, and in water inside plastic bags.
Besides loam and water, seeds were sown in Gibberellic Acid (GA3) and agar. Research in seeds has revealed that they have natural chemicals that control their growth. The Gibberellic Acid is a hormone that, among others, can break the seed dormancy, causing fast and uniform germination. It does not work for every seed, and the concentration required can be variable too. The concentration used was 914.8142 ppm (parts/million) in water, being the maximum suggested 2000 ppm. A higher concentration can cause an inhibition of growth instead of promotion, and then harmful effects to the plants later. Marc Hachadourian, from the New York Botanical Garden, has tried before to improve the germination rate with GA3 in the compost used, but he found it unnecessary.
Agar is a product obtained from red algae, and is useful as it is considered inert biologically. The concentration used was 8.5 g/L (Agar/Water), pH 5.8, being "a pH of 5.0 to 6.0 usually adequate. ( ) The minimum concentration used is about 6 g/L".
Normally, agar is used in a specific environment: sterilised surface, materials and hands. Because of lack of space in the Micropropagation Laboratory at Kew, the seeds had to be sown in an area that was not sterile, but as clean as possible. Because sugar was not a component in the agar, it was believed that there should not be any problem related to contamination.
An experiment with agar was done at Kew 2 years ago. Emma Fox, from the Palm House, and Dr. Tim Wilkinson, who worked in the Micropropagation Laboratory, extracted the embryo from 10 Victoria seeds and put them in agar + sugar to germinate. Only one grew, but the seedling died afterwards. Probably, the embryos were damaged during the process to take them off from the seed.
§ The seeds' viability:
There is also a divergence related to this factor. The Knotts say that the seeds do not remain viable for much more than one year, while at the National Botanic Gardens of Belgium and also at Kew, they keep their seeds for up to 10 years. Some seeds from 1996 have been tested too, sown in loam, in comparison with the seeds obtained from the USA.
Acknowledgements - References - About the author