Genetic Transformation And Biology Characteristics Of The Arabidopsis Thaliana Suspension Cells

Transformation of Arabidopsis thaliana suspension cells was performed. Different protocols and parameters affecting the transformation procedure were tested, and an optimized protocol was determined. Transformants was selected with antibiotic resistant gene screening and checked with histochemical GUS and PCR assay. Prior to transformation experiments, some relational biology characteristics of Arabidopsis thaliana suspension cells and Agrobacteria haboring certein plasmid were observed. The main works and results are shown as follows.Subculture Conditions and Growth Characteristics of Arabidopsis Suspension CultureThe experiments of Arabidopsis thaliana suspension culture were performed. Some subculture conditions were determined, and the growth characteristics of suspension culture was observed. The typical S-like growth curve in a subculture cycle was obtained. In normal condition, a logarithmic phase of growth of suspension biomass was occurred in 2-4 days after subculture. Than the growth of biomass slowed down quickly. However, its growth and time of logarithmic phase were changed with inoculation amount. 10ml/50ml of diluting ratio is advisable to subculture. The growth was slowed down even halted with 10ml/50ml of diluting ratio. Different hormones in media affected growth of suspension cells, but the result was unexpected. The biomass of suspension cells subcultured in B5 medium without any hormones was higher than with 0.1mg/L of BAP and 0.2mg/L NAA (as normal medium). Concentration of sucrose greatly affects growth of suspension cells. Changing it from normal 30g/L to 10,20,40 g/L, more or less inhibition of growth and viability were observed.The pattern of callus growth was observed. In general, each a 10 d of period is presented in callus induction phase, logarithmic growth phase and growth-stopped phase. Incubation temperature and humidity had a remarkable impact on both inducement and growth of callus. A certain surface humidity on plate and lower incubation temperature were indispensable.