A novel and efficient tissue culture method – "stem-disc dome culture" – for producing virus-free garlic (Allium sativum L.)

Abstract.

Using our previously reported stem-disc culture method as a basis, we have developed an efficient tissue culture method, "stem-disc dome (SD-Dome) culture", to eliminate viruses from infected garlic plants. Fifteen to 25 dome-shaped structures that formed on stem-disc explants from a single garlic clove were excised and maintained on phytohormone-free Linsmaier and Skoog medium, as in the usual shoot-tip culture. These excised dome-shaped structures grew independently into more than 5-cm-long shoots, rooted after 8 weeks, and were successfully transplanted to soil. The resulting plants showed no viral symptoms on their leaves, even when cloves of garlic plants with severe mosaic and yellow streak symptoms were used for the cultures. Examinations of virus infection in three generations of the progenies using the direct tissue blotting immunoassay and reverse transcription polymerase chain reaction showed that viruses were excluded during culture. Microscopic observations of transverse sections of tissues in different developmental stages during stem-disc culture – dome-shaped structures, shoot buds, and shoots – suggested that the elimination of viruses may be correlated with the stage of development and organization of vascular-bundle structures.