Circulated Culture System of Tobacco Hairy Roots for Rapid Evaluation of Viral Multiplication and Movement

From the hairy roots induced by an inoculation of Agrobacterium rhizogenes into tobacco (Nicotiana tabacum cv. Bright Yellow) leaf segments, we obtained the hairy root clone which could frequently regenerate shoots from calli produced at the basal sites of lateral roots branched from major hairy roots. The leaflets detached from the regenerated shoots produced secondarily the hairy roots and differentiated into the secondary shoots via calli in a similar manner. This circulatory culture system for the hairy root production and plant regeneration was available for an in vitro assay of multiplication and translocation of TMV. By the inoculation into detached leaves of the regenerated shoots with TMV, the virus-specific mosaic symptom was observed in the leaflets of the newly regenerated shoots, and the localization of TMV in the symptomless tissues such as stems, petioles, and secondary hairy roots of the infected regenerants was easily detected by direct tissue printing immunoblotting assay.