Plant cell and tissue culture together with the recent advances in recombinant DNA technology are getting great attention because these are the sources of innovation for the major agricultural industries and practice. However, despite the many successful applications of plant tissue culture in agriculture and horticulture, differentiation and especially plant regeneration, remain a major problem with a number of crop species. Relatively few crop species are capable of predictable regeneration. Even in those instances where regeneration has been reported, e. g. Nicotiana, Petunia and Arabidopsis, through empirical selection of explants and media, problems are often encountered with particular cultivars within a species. Hence, these problems constitute a serious block to crop improvement. Moreover, a detailed characterization of the competent host plant cell will be required if we are to further exploit the Agrobacterium system for transformation of recalcitrant plant species. In this review article we describe some possible opportunities which may lead to rational plant improvement.
High and wide range of concentration of sorbitol, 0.6M to 1.9M, was effective as osmoticum in combination with 2% each of Cellulase RS and Driselase, on the isolation of protoplasts from cotyledons of Avicennia marina and A. lanata. Protoplast yield varied from 106 to 107 cells per seed. Sorbitol at 1.3-1.4M was consistently effective of good yield.
The bacterial wilt resistant line LNSR-7 of tomato was isolated from self-pollinated progenies of leaf-callus derived regenerants by directly inoculating a bacterial wilt pathogen Pseudomonas solanacearum into injured roots of tested plants. The subsequent self-pollinated progenies of the line were examined for their fruit quality and resistance expression under natural cultivation conditions in a pathogen infested tomato field. During three generations of progenies, the tomato plants showing both the bacterial wilt resistance and the high fruit qualities comparable to the parental cultivar were selected in order to fix commercial characteristics of the line. The stable inheritance of the resistance in the subsequent self-pollinated progenies was further examined by directly inoculating the pathogen into the roots of test plants. Inoculated plants were planted in soil heavily infested with the pathogen to ensure exposure to the pathogen. Under these artificial inoculation conditions, the selected line was shown to be highly resistant to the disease. The resistance mechanism in the line was analyzed by examining multiplication and translocation of the pathogen in planta. The precise monitoring of infection behavior of the pathogen was successfully achieved using the genetically marked P. solanacearum. Consequently the present line LNSR-7 strictly limited secondary multiplication and translocation of the pathogen and suppressed the wilt induction by the pathogen.
To examine the mechanism for in planta detoxification of phytotoxicity of indole-3-propionic acid, the gene expression for glucosylation of this compound was detected in treated tomato seedlings by Northern and in situ hybridization with the specific probe. The probe was obtained by polymerase chain reaction of tomato chromosomal DNA using the primers designed on the basis of amino acid sequences which were highly conserved in several enzymes catalyzing glucosylation. The positive hybridization was intensively detected in stems of the treated tomato seedlings. The increase of hybridized transcript was well coincident with the accumulation of glucosyl indole-3-propionic acid analyzed by thin layer chromatography. The result suggests that the PCR clone obtained from tomato chromosome encodes partial sequences of a gene for glucosyl conjugation of the compound.