[ Rivercane Propagation ]
[ Seed Germination ]
[ Micropropagation ]
[ Macropropagation ]
Although canebrakes have now been recognized as critically endangered ecosystems ( 7 ,9), current restoration projects are few. Restoration efforts have been hindered by the limited supply of plant materials. Due to rivercane's sporadic flowering cycles (30-40 years), seed is difficult to obtain. Furthermore, rivercane seed, when produced, has low viability ( 7 ) and are recalcitrant ( 14 ), making for very low germination success. Although established culms propagate readily through rhizomes, transplantation of established culms is a labor-intensive restoration method, requiring 50-60 cm diameter clumps of soil containing 3-8 culms ( 15 ). Transplanting culms also have a low success rate due to embolisms which form when rhizomes are cut ( 29 ). Currently, researchers are investigating methods of vegetative propagation in order to increase the supply of plant materials for restoration projects. To date, most restoration projects involve the labor-intensive method of transplanting groups of culms from one area to another.
There is still little known about seed production in rivercane. So far, it appears that rivercane is self-incompatible, with few viable seeds produced in isolated patches ( 29 ). It also shows no indication of seed dormancy. During 2006 and 2007, rivercane flowering events were observed at many locations in the Southeast (Mississippi's western alluvial plain, Mississippi hill country, western Tennessee, western North Carolina, and central Alabama). With the availability of this seed, it has been possible to conduct germination research which may not be possible again for many years.
germpaper: germination paper
with rivercane seed
example of roll towel
replicate of roll
towels in clear box
Germination studies were conducted at Mississippi State University on seed from two populations of rivercane (Kituwah, near Bryson City and Caney Fork, near East Laport, North Carolina). Three germination methods were tested (roll towels, germination in clear boxes, and germination in dark boxes) with six temperature regimes ranging from 15-40°C (in 5° increments) ( 29 ). The Roll towel method used six reps of 100 seeds each of both populations. Seeds were placed on damp germination paper (2 on the bottom and 1 on top) and then rolled and placed in clear boxes. Germination between the two populations at low temperatures were similar, with both populations showing increased germination with increasing temperature regimes. Temperatures above 35°C, however, resulted in significantly lower germination rates. Optimal germination temperatures for the Caney Fork population was 30/20 and 35/25°C (day/night), with 58 and 56% germination, respectively. For the Kituwah population, optimal temperatures were 35/25, 30/20, and 25/15°C, with 28, 24, and 26% germination. Similar tests were conducted with unrolled germination paper in clear boxes, followed by the use of dark boxes in a following experiment. The roll towel method resulted in significantly higher germination, however, due to the recalcitrant nature of rivercane seed, additional studies of light versus dark were compromised. Seed was no longer viable after approximately 10 weeks at room temperature, making it impossible to follow up on the roll towel germination.
seedlings germinated from seed
Overall, seed viability was quite low, with 42% inviable seed from the Caney Fork population, and 72% from the Kituwah population. It is recommended that seeds be stored for no longer than 10 weeks at room temperature, though viability may be extended by refrigerating or freezing seed. Seed from the Caney Fork population was reduced by the presence of glumes surrounding the caryopsis.
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plant tissue culture
plant tissue culture
Micropropagation of rivercane has great potential in providing large amounts of plant materials available for restoration projects. Once established in culture, explants can be regularly divided to increase plant numbers without further need of source material. Micropropagation can provide rapid clonal multiplication with year-round production, which is not possible with other propagation techniques. Micropropagation techniques using tissue culture from newly developing rhizome and culms were investigated at the University of Memphis ( 29 ). Culms were 4-6 cm long and had at least 1 node and rhizomes were sectioned into 2-3 node segments. All leaves and sheaths were removed and the tissue was disinfected with a 70% ethanol solution. Different sizes of explant materials were tested to induce shoot initiation in sterile, laboratory conditions. The larger explants produced the greatest shoot growth, with no difference in survival rates. Shoot growth was also monitored in several growth media over time. At four weeks, both Murashige and Skoog (MS) and Woody Plant (WP) growth media with 1% agar produced the best growth. MS media is more commonly used in other bamboo propagation techniques.
Although these micropropagation techniques show great promise, results from this study were hampered by the lack of root formation. Further research is necessary to develop techniques to promote root development before this technique will be able to provide methods for propagating large numbers of shoots for restoration projects.
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Currently, macropropagation of rivercane shows the greatest immediate promise for providing enough plant materials for restoration projects. While the use of seedlings is ideal, the lack of consistent seed sources makes this unreliable. Seedlings also require 2-3 years of growth in order to provide rhizomes of significant size that can actively spread throughout an area. Macropropagation techniques are being studied at Southern Illinois University ( 30 ) and Mississippi State University ( 29 ). Researchers are working with propagation methods for both culm and rhizome propagation. Studies include: the collection of germplasm to avoid embolism, planting and harvesting rhizomes, the effect of light and moisture on propagule success, propagation medium, number of nodes (on culms and rhizomes), and planting methods.
Researchers at Southern Illinois University, Carbondale found the highest propagation success using larger rhizomes (10+ nodes) and exposing these rhizomes to daylight33. Rhizomes were maintained in a greenhouse under mist to prevent desiccation. They reported that rhizome success was largely influenced by the size of rhizomes and the collection date.
collected germplasms hanging
in polypipe tubing
Research at Mississippi State University ( 29 ) has led to a new technique for collecting plant materials while avoiding embolism of the culms. Researchers have found that by immediately placing the newly dug clump of culms into a large plastic tube and sealing both ends with about 1 liter of water, they can avoid embolism. They have found that polypipe, a large diameter (40-60 cm) white flexible plastic tube which is primarily used as temporary irrigation, works well because it is sold in 0.4 km rolls and can be cut to accommodate any height of culm. Once the culms are collected and sealed in the plastic tube with water, they are hung in deep shade for 6 weeks. Afterward, the plants can be planted in pots (half sand, half native soil). Using this technique, researchers have found a 96.3% success rate over all collected rivercane and a 100% success rate in 1-2 year old plants. Rivercane collected and brought to Mississippi State University is grown using a pot in pot technique commonly used in horticulture. The slightly larger socket pot is buried in the ground and the pot containing the rivercane culms is placed inside. This protects the roots from temperature extremes while allowing easy access to actively growing rhizomes.
rhizome: new growth
example of pot-in-pot
The harvesting of rhizomes for macropropagation should be done January through March, when plants shoot most vigorously. Research at Mississippi State University ( 29 ) investigated the used of plant growth regulators (PGRs) to enhance rhizome node activation and stimulate shooting. Experiments soaking different lengths of rhizomes in 1000 ppm BAP, GA4, and a combination of both resulted in the highest shooting rates using 2-3 node segments in a 60 minute soak of 1000 ppm BAP. In inactive nodes, this treatment resulted in 90% shooting. Nodes that were already active prior to treatments had 100% shooting with no treatment, 30 min in BAP and 60 min in BAP. Active nodes were negatively affected by the GA4 treatment. Rhizome segments with only one node had significantly lower shooting rates (4.7%), which were increased to 16.5% using the 60 minute BAP treatment.
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from rhizome segments
examples of active nodes (green),
dormant nodes (red),and dead node (black)
from culm segment
buried culms give
rise to new shoots