Production of sweet orange somaclones tolerant to citrus canker disease by in vitro mutagenesis with EMS

Xanthomonas citri subsp. citri (Xcc) is the causal
agent of citrus canker, a quarantine disease worldwide. This
disease is hard to be completely controlled by chemical
sprays, and the selection of citrus genotypes resistant to the
pathogen becomes one of the most important ways to control
the disease. The limited genotypes that have been found up to
now tolerant to Xcc are not commercially cultivated. Sweet
orange [Citrus sinensis (L.) Osbeck] is one of the most
important citrus species widely cultivated in world and all the
orange cultivars are susceptible to canker disease. Therefore,
this study aimsat the establishment of an efficient protocol for
in vitro ethyl methane sulphonate (EMS)mutagenesis and the
selection of ‘Bingtang’ sweet orange somaclones tolerant to
citrus canker disease.Mutation was introduced by treating the
cell suspension of embryogenic calluswith 1.5 %ofEMS for
1 h (the lethal concentration). A crude extract produced from
the pathogen solution was used as the selection agent. Sweet
orange leaves inoculated by Xcc-crude extract showed the
symptoms similar to those inoculated by Xcc bacterial inoculum.The young shoots of susceptive genotypes cultured
in 10 %of Xcc-crude extract solution becomebrown and died
in 1 week, while the resistant citron C-05 grew normally
under the same conditions. After two steps (cell suspension
and plants) of selection by Xcc-crude extract solution, the
survival plants were tested by in vitro and in vivo inoculation
with Xcc. One somaclone, named DG-2, was identified to be
resistant to the canker disease. The results suggested that the
established protocol for somaclones variation by treating
sweet orange callus with EMS and in vitro selection of the
somaclones tolerant to canker disease by the pathogen crude
extract was effective. The gained swee