Evaluation of a Simple Automated Bioreactor for the Production of Pelargoniums Progress Report –September 1992
Date 28 August 1992
Title of Project Evaluation of a Simple Automated Bioreactor for the Production of Pelargoniums
Institution where work is being conducted The Pennsylvania State University
Amount of Endowment Grant $ 5, 000
Covering Period 6/92 to 5/93
Anticipated Date of Project Completion/Final Report 6/96
Individual(s) Conducting Project:
(List Project Leader First)
Richard N. Arteca - Title Professor
Telephone Number 814-863-2252
Evaluation of a Simple Automated Bioreactor for the Production of Pelargoniums
Richard N. Arteca
Pennsylvania State University
- A. Project Objectives:
- Our major objectives for the first year of this project are to produce callus toPelargonium leaves, maximize growth rates while maintaining chromosome stability, to
begin to develop methods for cryopreservation of cells and begin evaluating different
hormone treatments to induce the regeneration of plantlets.
- B. Summary of Work Since Project’s Inception Suitable for Dissemination:
- At the present time the results to date are preliminary and not suitable for dissemination.
- C. Results to Date:
- During this first year we had to modify several parameters initially proposed. First, weshortened the surface sterilization procedures initially proposed because they were harmful
to the young succulent tissue which we used. Second, we have found that Musashige and
Skoog basal salt medium caused surfaced sterilized Pelargonium tissue to turn black, exude
a black colored substance into the agar and died. This problem was overcome by switching
to Gamborg’s B5 medium which contains lower salts and has been used for all subsequent
experiments. After overcoming our initial problems we were able to generate callus from
Pelargonium x hortorum cv. Veronica in the light and dark after several months in culture.
Callus proliferation was found to be faster in the dark than in the light. With these
problems taken care of we can now easily produce additional callus and suspension culture
cell lines if necessary in the future. We have expanded several callus cell lines 3 initiated in
the light and 3 initiated in the dark for further experiments and these cell lines will be used
for future experiments unless complications arise. We have used one of the callus cell lines
to produce a cell suspension culture line with moderate growth rates and are presently
expanding out this cell line for further experiments. Each of the cell lines have maintained
chromosome stability thus far. We have thus far been unsuccessful in cryopreserving
either callus or suspension culture lines, however, we will be using different
cryoprotectants to overcome this problem. We are currently modifying the growth media
with different components to maximize the growth rates of our callus and cell suspension
culture lines. In addition, we have begun experiments with different ratios of plant
hormones in order to promote shoot formation. Using the proper ratios of
benzylaminopurine and naphthalene acetic acid we have had some success inducing shoot
formation and are presently waiting for these shoots (see attached photo’s) to enlarge prior
to transferring to another hormone regime to initiate roots and finally transfer to the
greenhouse.
- D. Future Plans Covered by the Endowment Grant:
- Our goals for the remainder of the 1991 to 1992 research year are to begin manipulatingconditions to maximize the growth rates, shoot and root formation in our callus lines and to
use different cryoprotectants in order to cryopreserve our existing cell lines. The proposed
experiments for the 1992-1993 research year are to do the following:
- 1. Manipulate environmental conditions, plant hormone amounts and ratios and othermedia supplements to promote maximum growth of callus and cell suspension growth;
- 2. Induce plantlet formation and maximize the number of plantlets formed withdifferent hormonal treatments in callus and cell suspension cultures;
- 3. Once we have maximized plantlet formation we will transfer these plantlets togrowth media containing different ratios of plant hormones in order to induce root
formation and then grow the plants in tissue culture to increase the size and number. We
will then begin work to acclimate these plants for transfer to the greenhouse where they will
be grown to flowering.
- It should be noted that each of the proposed experiments will depend on the rate atwhich the plants grow prior to moving to the next step, however, the entire project should
be successfully completed within the proposed four year period. At the present time we are
keeping up with the schedule initially proposed.
- After these goals are achieved we will evaluate bioreactor technology for the productionof Pelargonium plants. This will be accomplished by seeding an airlift bioreactor with cells
grown in suspension culture and do the following:
- - treat with hormones to promote maximum growth of callus;
- - induce plantlet production with treatments developed earlier and transfer plantlets tosoil and grow.
- The last phase of this project will be to design a larger bioreactor and evaluate how initialstudies work on a commercial scale.
- E. Anticipated Benefits for Floral Industry:
- The overall goal of the proposed research is to develop an automated bioreactor whichcan be used to increase the commercial production of selected Pelargoniums with desired
characteristics such as new genotypes or disease-free plants rapidly and efficiently. The
availability of such methods will also lead to increased branding and selection efforts which
in turn will promote increased quality of Pelargoniums at a reduced cost which is a true
benefit to the floral industry.
