Preliminary results for Monarda bradburiana germination after 30 days

March 2, 2015 Alejandra Feliciano

Two of the three species in Group A have completed their first 30 days in the growth chamber and have been moved to the greenhouse.

Group A species received no moist-cold stratification treatment. They were placed in the growth chamber at 24C immediately after imbibement. Preliminary results for this treatment of the three species is listed on the table below. Although the percent germination for Vernonia lettermannii is very low, it is higher than anticipated considering the 2% viability determined by the preliminary TZ test.

Monarda bradburiana had significantly higher germination rates than the other two species, which makes it a more likely candidate for seed mix production. Stratification treatments are already proving to improve germination rates for this species as well, with germination of 46% after 3 days in the growth chamber.

Monarda bradburiana under no stratification treatment. Top shows first day of germination; bottom image shows growth after 30 days in the growth chamber. — *Monarda bradburiana* under no stratification treatment. Top shows first day of germination; bottom image shows growth after 30 days in the growth chamber.

Y-axis shows percent germination. X-axis shows days in growth chamber. Liatris microcephela is still in the growth chamber and data collection is ongoing. — Y-axis shows percent germination. X-axis shows days in growth chamber. *Liatris microcephela* is still in the growth chamber and data collection is ongoing.

Preliminary results. Note Liatris microcephela data collection is ongoing. — Preliminary results. Note *Liatris microcephela* data collection is ongoing.

Seeding Monarda bradburiana

January 22, 2015 Alejandra Feliciano

Three treatments of Monarda bradburiana will be documented for this study. All treatments were imbibed and seeded on January 22, 2015 into 3 x 4 x 18 perforated Jiffy trays filled with SunGro Sunshine SB300 Universal professional growing mix. Seeds were sown in 10 rows of 10 seeds each, for a total 100 seeds per treatment. Treatment A was placed in a growing chamber at 25C on the date of imbibement. Treatments B and C were placed in a cooler at 5C on the day of imbibement.

Treatment A: 0 - day moist-cold stratification. Placed in warm growth chamber immediately after imbibement.
Treatment B: 30-day moist-cold stratification. Placed in cooler immediately after imbibement.
Treatment C: 60-day moist-cold stratification. Placed in cooler immediately after imbibement.

Seeding tray: 3" x 4"x 18" perforated plastic tray. — Seeding tray: 3" x 4"x 18" perforated plastic tray.

Water spraying nozzle (bottom) used to imbibe seed.

From background to foreground: Treatment A, B, and C, respectively.

Tetrazolium (TZ) Testing

January 22, 2015 Alejandra Feliciano

After the seed conditioning phase was complete, I proceeded to test the seed viability. Tetrazolium tests are performed at the ISU Seed Testing Lab. In this test, seeds are imbibed overnight, then pierced the following day and soaked in a tetrazolium solution. The solution is then absorbed through the piercing, tinting the seed.

Because of the limited number of seed available for the project, two replications of 50 seeds were performed for each species in lieu of the typical 100 seeds per replication.

M. bradburiana TZ test results. — *M. bradburiana* TZ test results.

V. lettermannii TZ test results. — *V. lettermannii* TZ test results.

Preliminary results were reported on January 23, 2015. The official report was published on January 26, 2015. The results were as follows:

M. bradburiana: 72% viable seed reported.
V. lettermannii: 2% viable seed reported.

The cause for poor embryo development in this batch of V. lettermanni is unknown. However, due to the low viability of V. lettermannii, a replacement species must be used for this study.

Conditioning Vernonia lettermannii

January 21, 2015 Alejandra Feliciano

Vernonia lettermannii presented a conditioning challenge in the threshing stage. Threshing is the process by which the seed is mechanically separated from its plant parts, in this case a shell and coma. After multiple failed attempts to separate comas from seed, we found the most effective to be gently rubbing with a sponge against an elongated slotted screen. This produced shells, seeds, and a kind of wool (derived from the comas).

V. lettermannii prior to threshing. The beautiful maroon-colored comas were difficult to remove.

Threshing by gently rubbing against a slotted screen produced a wool-like material from the coma debris.

'Ironwool' is a by-product from the ironweed threshing process.

V. lettermannii seed can be seen here caught suspended vertically on the screen. — *V. lettermannii* seed can be seen here caught suspended vertically on the screen.

Separating threshed debris from seed using a pressurized air nozzle and varying screen sizes.

Once separated, the seed could easily become trapped within the 'Ironwool' material, becoming nearly impossible to separate. For this reason, special care was taken during conditioning to minimize contact between both substances.

Conditioning Process

Thresh collected seed by rubbing with sponge against screen.
Use pressurized air nozzle to separate threshed debris from seed by blowing the material through a series of decreasing size screens.
Separate poorly filled seed from heavier seed using South Dakota Air Blower.

Many thanks to Alan Gaul for his help in the conditioning process.

Conditioning Monarda bradburiana

January 20, 2015 Alejandra Feliciano

The first step in this germination study was seed conditioning. Seed conditioning refers to all treatments required to produce clean, quality seed. Monarda bradburiana seed were collected from Dr. Denny Schrock's private garden and brought to the ISU Seed Science Center for conditioning. The seed itself is very small--about 1mm in length. This meant that bulk conditioning equipment would not be usable for the small batch. The conditioning process used proceeded as follows.

M. bradburiana seed prior to shaking the sieve. some debris visible. — *M. bradburiana* seed prior to shaking the sieve. some debris visible.

Metal sieves used to separate large debris from M. bradburiana seed. — Metal sieves used to separate large debris from *M. bradburiana* seed.

South Dakota Seed Blower residue (seen here) collects lighter weight particles in angled shelves at the top of the acrylic blower tube. In this case, it has collected light-weight debris and potentially damaged light-weight seed.

Makeshift vibratory deck separator (design by Alan Gaul). Easily-rolling objects, such as round seed, collect down into the lower blue tray while remaining objects vibrate upwards over the plate. — Makeshift vibratory deck separator (design by Alan Gaul). Easily-rolling objects, such as round seed, collect down into the lower blue tray while remaining objects vibrate upwards over the plate.

Magnifying glass station used for hand-sorting and seed counting.

Deatiled view of M. bradburiana seed on magnifying glass station. — Deatiled view of *M. bradburiana* seed on magnifying glass station.

Conditioning Process

Sifting through metal sieve separates larger debris from seed. Most effective was 0.838 cylinder.
Vibratory deck separator separates rolling seed from non-rolling debris.
South Dakota Seed Blower isolates light-weight seed and debris. This seed has presumably not filled-in properly and is unlikely to have fully developed embryos.
Hand sorting using magnifying glass station and tweezers.

Many thanks to Usha Arora and Alan Gaul for their assistance in this process.