The Study On Transposon Activition In Long-term Subcultured Citrus Calli

Callus is regard as an alternative material for the germplasm preservation because of its features of high speed for reproduction, convenience for conservation and little space occupation. However, there are kinds of somaclonal variation during the long-term subculture. The transposon activation is consented as one of the molecular mechanisms to explain the somaclonal variation. What‘s more, studies recently claim that the transposon elements(TEs) are probably involved in the evolution of genome, the regulation of gene expression, response to the stress and other process. The study on fruits‘ transposon activation is a great of significance for keeping the stability of the genetic material during the germplasm preservation, finding new ways for germplasm innovation and providing new ideas for understanding the bud mutation. Our study detected the somatic embryogenetic ability of the Valencia callus after long-term subculture, and we analyzed the transposon activation of the long-term cultured Valencia callus through several method including Transposon Display(TD), Southern blot and real-time PCR. Meanwhile, we explored the factors that have influence on the transposon activation. The main results list as follows:1. We detected nine lines of Valencia callus. Six of them are able to produce the embryoid. The ploidy of the embryoid is as same as the callus which produced the embryoid. However, only one line of Valencia callus whose embryoid is able to grow into seedling. So we conclude that the long-term subcultured Valencia callus maintains the somatic embryogenetic ability but that ability reduces during the long-term subculture.2. The TD results indicate that the haplochromosome copy number of Csgypsy1,Csgypsy2, Cscopia3, Cs Mu DR1, Cs MITE1, Cs Tcs1, Cscopia1, Csh AT1 increased in the long-term cultured Valencia callus, compared with the newly induced Valencia callus. And the haplochromosome copy number of Csgypsy1 and Csgypsy2 in the polyploidy Valencia callus is higher than the haploid or diploid ones. This conclusion is also proved in the Southern blot of Cscopia1. At the same time, the results of Southern blot digested by the DNA methylation sensitive restrict enzymes verify that there exists the DNA methylation at a certain extent, in the TEs areas.3. The transcriptional expression of Csgypsy2, Cscopia3, Cs Mu DR1, Cs MITE1 are at low level overall, while Csgypsy1 is highly expressed. Because of the low temperature stress, the transcriptional expression level of Cs Gypsy1 and Cscopia3 increased distinctly. The copy number of Cscopia3 increased markedly as the low temperature treatment time prolonged. In a word, low temperature stress is one of the factors that inducing the transposon activation in citrus callus.