Research On Plant Tissue Culture And Expression Vector Construction Of Drought And Salinity Induced Genes In Alternanthera Philoxeroides

Drought is the most severe abiotic stress to crops, and so is it with salinity stress which causes damages to plants by ion toxicity and drought similar osmotic stress. Damages of drought and salinity to plant growth and crop productivity exceed the sum of the other natural disasters. Compared with traditional methods, identification and genetic manipulation of drought and salinity related genes to crops are efficient ways to stress resistance. Alternanthera philoxeroides (Mart.) Griseb. is a kind of plant with strong drought and salinity adaptation, thus it would be a good material for researching the mechanism of plant stress tolerance, as well as a resource for those stress resistance related genes. Besides, its environmental decontamination ability and medicinal value appealed to more and more people.In this paper, our research includes two parts. One is on the plant tissue culture in Alternanthera philoxeroides. The effects of culture factors, including plant growth regulators, explants types and initial light conditions, on callus induction and shoot regeneration were examined by orthogonal tests. The results showed callus induction was significantly stimulated by 2, 4-D, followed by explants types and duration of culturing in the dark; and shoot differentiation was affiliated by initial continuous light and 6-BA. The optimum conditions for callus induction were to use stem explants cultured on semi solid MS supplemented with 6-BA (0.5 mg l-1), 2, 4-D (1.0 mg l-1) with an initial dark period of 20 days;while for shoot regeneration, the best results were obtained using calli cultured on semi solid MS supplemented with 6-BA (2.0 mg l-1), culture in continuous light. All media were supplemented with sucrose (30 g l-1), and maintained at 25±2℃. This in vitro regeneration system would contribute to the molecular mechanism research of drought and salinity tolerance as well as genetic manipulation in Alternanthera philoxeroides.The other part is on expression vectors construction for gene sequences S2910 (the accession number: DQ492290.1) and D2714 (the accession number: DQ985704.1), which were both induced to be expressed in Alternanthera philoxeroides during drought and salinity stress. S2910 is an EST sequence,for which we constructed an anti-sense expression vector named pBI121-S2910 with plasmid pBI121. D2714 is a full length of gene sequence; an expression vector named pGAPZα-D2714 was constructed by cloning the ORF of D2714 gene to the vector pGAPZαA, which could be expressed in Pichia pastoris. Construction of expression vectors is pre-condition to investigate the function of the sequences, and would finally contribute to genes selection of stress tolerant.