Studies On The Tissue Culture And Gene Transformation Of Polar 84K

With the leaves as explants, the tissue culture and plantlet regeneration system of polar 84 K were established and the factors influencing plantlet regeneration and the hyperhydric shoot of polar 84 K were investigated. And a further study was made on gene transformation of polar 84 K with HVA1.( ) The research on the tissue culture and plantlet regeneration system of polar 84 KWith the leaves as explants, the effects of different concentrations and types of hormones on the dedifferentiation and redifferentiation of polar 84 K were investigated. The result showed that callus could be obtained after seven days of culture on the MS basal medium supplemented with ZT 0.5mg/L N 2, 4-D 0.5 mg/L^ IAA0.5 mg/L and NAA 0.5 mg/L. The suitable medium for bud formation was MS basal medium supplemented with ZT 1.0 mg/L and NAA 0.1 mg/L. On the suitable medium adventitious buds could be induced after 15 days of culture and the induction rate was 100%. The number of buds on each leaf ranged from ten to twenty. The plantlet could be got when the adventitious bud was 2 cm to 3 cm in height, and were excised ,then continually cultured on the medium (1/2 MS supplemented with IB A 0.2 mg/L) for ten days.( ) The research on the hyperhydric shoot formation of polar 84KIn the course of the tissue culture and plantlet regeneration of polar 84K, there were some hyperhydric shoots. In order to reduce the count of the hyperhydric shoot, an investigation was made on the effects of number of shoot inoculated per vessel and the concentration of hormone (6-BA, NAA), sucrose, agar, macro nutrient on hyperhydric shoot formation of polar 84K. The result showed that with the increase of the concentration of agar, the hyperhydric shoot formation rate of 84K polar obviously reduced. Considering the proliferation rate, we thought the suitable concentration of agar should be 7g/L. As to the count of shoot inoculated per vessel, the more shoots were inoculated in each glass jar the more hyperhydric shoots formed. For reducing hyperhydric shoot formation rate, it was better to inoculate two shoots per vessel. No obvious relation was found between the hyperhydric shoots and the concentration ofhormone, but the effect of concentration of hormone (6-BA, NAA) on proliferation rate of polar 84K was obvious. We drawed the conclusion that high concentration of sucrose could prevent from formatting hyperhydric shoot, but influence proliferation rate and normal growth of polar 84K at the same time. In the experiment of the impact of the concentration of macro nutrient on formation hyperhydric shoot of polar 84K, with the increase of concentration of macro nutrient, hyperhydric shoot of polar 84K became more. Neither too high nor too low concentration of macro nutrient was good for the growth of polar 84K.On the condition that a large number of plantlets could be obtained and the number of hyperhydric shoots could be fewer, the suitable medium for proliferation of poplar 84K was MS basal medium supplemented with 0.5 mg/L 6-BA 0.5mg/L NAA, 30g/L sucrose, 7g/L agar and two shoots were inoculate in each vessel.( ) The research on the drought resistant gene( HVA1) transforming poplar 84KWith the leaves as the transform receptor system, drought resistant gene (HVA1) was transformed into poplar 84K by Agrobacterium and Kanamycin resistant (Kmr) plantlets were obtained. In the course of gene transformation, the concentration of bacteria, the time of immersion, co-culture and the concentration of antibiotic were studied and the result showed: the suitable factors were 10-15 minutes immersion, 2-3 days co-culture, ceftriaxone 200 mg/L and kana mycin 30 mg/L in the selection medium.The research on high frequency transformation system and gene(HVAl) transformation of polar 84 K not only supplied a new experimental system for polar 84K in many foundational theory studies including physiologic development, cell differentiation and morphogenesis etc, but also laid a foundation for the improving breed, acceleration of breeding and rapid propagation o...