The Research On Occurrence Of Transgenic Cherry Tomato Mutant With HBsAg Gene

In this experiment, the new plant expression vector pCAMBIA1301/HB and pCAMBIA1301were transformed into cherry tomato explants by Agrobacterium LBA4404. A series of transgenic plants were successfully constructed. And two groups of transgenic cherry tomatoes were taken varient analysis. Specific experimental progress are as follows:1The explants of cherry tomato could regenerate on the culture medium of MS+6-BA3.0mg/L+IAA0.2mg/L, and the number of regenerated shoots on each explant and the budding rate can reach the maximum.2Transformation of hepatitis B(HBsAg) gene and pCAMBIA1301vector into cherry tomato cotyledon explants by Agrobacterium was carried out. After PCR and southern blot,22transgenic plants with HBsAg and6transgenic plants with empty vetor were constructed.3Based on cytological analysis, the number of stomatal guard cell in the22different strains of transgenic cherry tomatoes with HBsAg has not obvious difference with the control plant. The average number of chloroplast of them and the amplitude of the number are basic similar. Flow cytometry analysis results show that the plants are diploid or near diploid. In addition, calculations of chloroplast’s number in stomatal guard cell of cherry tomatoes with empty vetor show that, there is little difference between the transgenic and control plants. There is great difference between the No.4strain and control plant on the average number of chloroplast. But the ploidy test of DNA showed that, all the transgenic plants with the empty vector are diploid or near diploid. There is no variation on ploidy of DNA.4Through the detection of pysiological indicators, the N244plant of transgenic cherry tomato with HBsAg has a significant variation with the control plant. There is no great difference between the rest of transgenic plants and the control one. Compared with control plants, only the No.3plant of transgenic tomatoes with empty vector has a larger variation on the physiological indicators. And their resistance is weak.5Comprehensive analysis shows that, all the transgenic cherry tomatoes have no ploidy variation. Therefore, the insert of the HBsAg gene does not lead to the produce of mutation inevitable. Previous studies in our laboratory have ruled out the possibility of somatic cell clonal variation. The external pressure in the transgenic processes may increase the mutation rate of plants. So the emergence of the mutant may be related to the process of genetic transformation. In addition, in the calculations of the chloroplast in stomatal guard cell and the experiments of the physiological indicators, there are a few variations in some plants compared with the control plants. The reason of this variation will be a further research.