Cryopreservation System Of Pummelo Germplasm Resources

It's the traditional method for the conservation of plant germplasm resources to plant in the open fields, which is limited its application on the long-term storage of germplasm resources because of effects from natural climate, space limitation, lack of managerial cost, and other reasons. Recent years developed from tissue culture technique, LN cryopreservation was proved to be an ideal way to conserve plant resources due to its advantages, such as conserving stability, high efficience, less expense and so on nowadays. In order to construct its technological system of cryoconservation and provide reference for the long-term and stable storage of pummelo resources and other fruit germplasms, we began to study the cryopreservation by vitrification of pummelo from the Western of Hubei Province. And the main results are as follow:1. Approaches for in vitro culture of mature pummelo stems were conducted. According to the growing principle of the fungi spores and its different growing stage s sensitivity to fungicide, the experiment obtained an effective disinfecting way with the materials of mature pummelo stems of about August: stems,3~4cm in length, were disinfected with 70% alcohol 30s,and 0.1% HgCl2 solution 20min.Then the explants were kept for 2d and repeated disfected with 0.1% HgCl2 solution 20min.After washed with distilled water, the explants were cut into small stems,0.5cm long with one lateral bud, and cultured in the primary medium. With the disinfection way, the contaminated rate of explants greatly decreased, and got the successful rate 85.6%.The induced medium of lateral buds was selected, the results showed that the optimal medium was MT+BA2.0 mg/L+GA30.5mg/L+NAA0.1mg/L+glucrose30g/L.The new branches were sub- cultured when 1.0~ 1.5cm long, growed well, and no callus was observed.2. The basic conditions for pummelo shoot-tips cryopreserved by vitrification were studied. With the high concentrated osmoprotectant of sucrose, preculture with Jingyang-baiyou Pummelo, obtained the optimal precultural pattern, 0.3M (ld)+ 0.5M (ld)+ 0.7M (Id), was highly efficient and the survival percentage 81.4%.Disiccation time and temperature of PVS2 were studied with the material Yichang Bendiyou Pummelo No.4,the results showed the recovery rate was relatively high, 72.2%,when the material was dehydrated 20min at a temperature of 0℃, and better than the results of the treatment conducted under the temperature 20℃. Effects of size of shoot-tips of 4 cultivars were researched. The average survival rate of shoot-tips with size 2.0~2.5mm,was higher than those of the other two sizes(1.0~1.5mm,3.0~3.5mm),and which was 60.6%.And the optimal rooting medium l/2MT+NAA0.5mg/L+Sucrose40g/L, rooting percentage 90.6%.