Application Of In Planta Callus Induction And Bud Regeneration System In Chinese Jujube

Chinese jujube (Ziziphus jujuba Mill.) ranks first in all dry fruit trees in terms ofproduction and its industry developed rapidly, which urgently require elite cultivars.However, it is difficult to conduct traditional cross breeding, and the selection breeding isconsidered time-consuming. Therefore, people start to explore other breeding methods,such as polyploid breeding etc. Currently, we had developed in vivo callus induction andbud regeneration technique on jujube and hereby achieved multiple chimera-freepolyploidy Chinese jujube and sour jujube germplasms. Here, we try to carry out in plantatransformation, EMS mutagenesis and polyploid breeding in jujube based on in vivo callusinduction and bud regeneration technique to lay a foundation of transgenic breeding,mutation breeding and polyploid breeding. The results were shown as follows:1. Branch diameter showed effects on bud regeneration. The result show that thenumber of the regenerated buds increased with the diameter of the branches. The branchesin diameter of2.0-2.5cm produced maximum buds, however, in consideration of numberof branches in large diameter and its damage to trees, the branches in diameter of1.5-2.5cm were most suitable.2. Infection period of agrobacterium rhizogenes influenced callus growth, budregeneration and genetic transformation rate in ‘Dalilongzao’. Infection were conducted at1st,4th,7thor10th day after branches being cut respectively and the results showed that thecallus was inhibited during the early days until20thday. The number of buds analysisshowed that the most suitable infection period was before7thdays. However, notransformated buds were acquired.3. Concentration of Agrobacterium tumefaciens solution influenced callus growth,bud regeneration and genetic transformation rate in ‘Yueguangzao’. Three concentrations(OD600=0.4,0.8,1.2) of Agrobacterium tumefaciens solution showed no effect on thecallus growth. In the20thday, we found that the concentration of the Agrobacteriuminfluenced the buds regeneration. We achieved maximum buds when the concentration was0.8. However in the30thday, the number of the buds had no significant difference. So we inferred that the different concentration of the Agrobacterium had influence on budregeneration only in early days. We achieved two transformation buds when theconcentration of the Agrobacterium was0.4which was perceived as the suitableconcentration. The two transformation buds were observed grown slowly and weak whichmight be due to the influence of Agrobacterium.4. Infection time showed no effects on callus growth, bud regeneration and genetictransformation rate in ‘Yueguangzao’. Callus grade and bud number showed nosignificantly difference when the infection time was12h,24h,36h or48h.5. Infection times influenced callus growth, bud regeneration and genetictransformation rate in ‘Yueguangzao’. At the10thday after branches being cut, the callusgrade which was infected three times was lower than those of two times or once infectionand control. At the20thday after branches being cut, though the difference of callus gradesamong different infection times was not significant, it also showed that the number of theregeneration buds decreased with the increase of the infection times. The number of theregenerated buds which were infected once showed no difference with those of control butwas significant higher than those with two or three times infection. However, in the30thday after branches being cut, the number of regenerated buds among infection times had nosignificant difference. It is a pity that no transformation buds were achieved. Inconsideration of both bud number and time consuming, one time infection was suitable.6. Elite table Chinese jujube cultivars including ‘Wuhefeng’,‘Jingzao39’,‘Lengbaiyu’ and ‘Liuyuexian’ were treated with colchicine based on in vivo callusinduction and bud regeneration technique. And finally, triploid and chimera of diploid andtetraploid were achieved in ‘Jingzao39’.7. EMS mutation (with concentrations of0.1%、0.3%、0.5%、0.7%and1.0%) in‘Yueguangzao’ through in vivo callus induction and bud regeneration technique were alsoconducted. The results show that both the phosphate buffer (PBS) and EMS solution wereharmful to callus growth and bud regeneration. The regeneration buds need furtherobservation to be sure weather mutation occurred or not.