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Development
of Disease Resistant Japanese Flowering Cherries Using Multiple Genes
Roger
J. Sauve, Jing Tian Ling and Christopher J. Catanzaro
Cooperative
Agricultural Research Program
Seminar Series
Tennessee State University, Nashville, TN
March 31,
1999
Introduction. Japanese flowering cherry trees are
of increasing importance in residential, recreational, public and
industrial plantings. However, diseases that affect cherries are
numerous. In spite of these diseases, demand for these plants continues
to increase. Bacterial canker caused by Pseudomonas syringae pv.
syringae is the single most important factor limiting the successful
cultivation of Japanese flowering cherries in Tennessee. Incidence of
this disease has caused substantial losses to most Tennessee growers
that have attempted to grow this tree.
Objectives. In this project, we propose to
transform Japanese flowering cherries with genes to resist pathogens.
Methodology.
Step One: development of a
regeneration system. Seeds of several species of Japanese flowering
cherries have been obtained. Prior to germination, the seed coat was
removed and seeds were disinfected by immersion in a 1.5% (v/v) sodium
hypochlorite solution containing 0.1% (v/v) Tween-20 for 20 min.,
followed by three rinses with sterile water. Disinfected seeds were than
placed in 10 x 15 mm test tubes that contained 0.8% agar for
germination. Uniform-sized hypocotyls will be selected and cut into 1 cm
segments. Once a suitable procedure is developed for seed germination,
the hypocotyl explants will be plated on a basal medium that contains
inorganic and organic salts of Murashige & Skoog (1962) and 30 g/l
sucrose. Depending on the experiment, the basal medium will be
supplemented with different concentrations of thidiazuron (TDZ),
benzyladenine (BA), indole butyric acid (IBA) or gibberellic acid (GA).
All media will be adjusted to a pH of 5.7 with 1 N NaOH and solidified
with 8 g/l agar (Sigma) prior to autoclaving. Growth regulators,
sterilized through 0.22 µm filters, will be added to the media after
autoclaving. All media will be dispensed as 15-ml aliquots into 15 x 90
mm Petri dishes. Step Two: genetic engineering. Chimeric genes
encoding for compounds that are toxic to fungal and bacterial diseases
of cherries will be introduced into commercial varieties and elite
breeding lines by particle bombardment. Step Three: evaluation.
Field assessment will be conducted to select outstanding individuals for
incorporation into a breeding/varietal release program.
Results. We are in the initial stages of
research. The first step for the genetic engineering of any plants is
the development of a regeneration system. Thus far, we have obtained
seeds of several Japanese flowering cherries and are currently
evaluating methods for the efficient germination of seeds. We have
obtained a limited amount of seed germination through the use of
stratification, acid treatments, and seed coat removal. Once we develop
an efficient germination system, we will use hypocotyl segments as
explants to develop a regeneration system.
Impact. The U.S. nursery industry is significant
and represents a high growth sector in U.S. agriculture. Ornamental
cherries would gain a measurable benefit from the introduction of genes
conferring horticulturally valuable traits.
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