Effects of Agrobacterium tumefaciens on the Symptoms of Paulownia sp. Plantlet in Vitro Cultured

By using Agrobacterium tumefaciens isolated from poplar crown gall disease with the hormone-producing genes in the T-DNA to inoculate healthy and infected Paulownia plantlets with phytoplasma, it is showed that tumorigensis of diseased plantlets dropped apparently and the symptoms of witches'broom suppressed to some extent. The T-DNA was transformed into Paulownia resulting in tumor formation independent of exogenous hormone addition and keeping subculture of tumor tissues for more than 2 years, thus confirming that the tumor tissues gained the ability to synthesize cytokinin and auxin by itself. Based on the conserved sequence of isopentenyl adenosine transferase gene(ipt) of Agrobacterium tumefaciens Opine pTil 5955 strain, a pair of DNA primers(CYT and CYT′) were designed and synthesed. A 427 bp polymerase chain reaction(PCR) product, the same size as the ipt gene fragment of pTil 5955 was amplified in association with Agrobacterium tumefaciens strain causing poplar stem gall disease rather than recombinant plasmid without this gene. The specific fragment of 427 bp was also amplified using the DNA extracts from transformed tumor tissues of two Paulownia clones(AT-ZH and AT-T35) with the Agrobacterium tumefaciens as templates, therefore further verifying the insert of T-DNA to the chromosome DNA of Paulownia and Paulownia can be surely transformed through Agrobacterium tumefaciens Ti plasmid vector. The specific 427 bp product was not amplified by using total DNA extracts of both in vitro cultured healthy and infected paulownia and sweetpotato infected with phytoplasmas as templates. When the transformed tumor tissues were grafted onto the infected in vitro cultured Paulownia plantlets with phytoplasmas, the symptoms of witches'broom reduced apparently, including reduced severity, prolonged survival time and increased rooting ability of the plantlet, which, on other aspect, suggests the involvement of the hormone metabolism in paulownia-phytoplasma interaction.