Effects Of CPPU On Tissue Culture Of HASKAOP And Germplasm Resources Of Cryopreservation

The Japanese Hokkaido special product High nutrition small fruit tree(Lonicera caerulea L.var.emphyllacalyx Nakai) which originally produces to the Japanese Hokkaido is an important fruit indigenous.The fruit has always been recognized for its nutritional and medicinal values.In order to successfully preserved the germplasm resources of HASKAOP,this test applied a new cytokinin CPPU which was found in recent years to mall fruit trees in tissue culture,explore the best use of the concentration of promoting propagation of small fruit tree,and used the method of cryopreserved to preserved germplasm resources,with a view to provide a theoretical basis for the large number of in vitro preservation of small fruit tree germplasm resources.The main results of this research are as follows:1.In order to explore the effects of CPPU on hokkaido specialty little fruit tree in the application of tissue culture,we carried out a test of CPPU on alone and after the combination of 6-BA in hokkaido specialty little fruit tree internode differentiation of adventitious buds's induced effects.The results showed,CPPU alone add the best-induced concentration is 0.2 mg/L,a combination of6-BA 0.5mg/L+CPPU 0.5 mg /L was the best formula for hokkaido specialty little fruit tree internode differentiation of adventitious buds,besides,the effect of CPPU is better than 6-BA.2.Internodes of hokkaido specialty little fruit tree were successfully cryopreserved with the vitrification technique.A suitabl(?) procedure was established as follows:internodes about 0.5～1.0cm in length were precultured for 2 days on MS medium supplemented with 0.7mol/L sucrose.Dissected internodes about 2.0～2.5mm in length,pretreated in a solution of 60%PVS2 for 20～30min at room temperature,and then treated with solution of 100%PVS2 at 0℃for 60 min.After changing with fresh PVS2,intemodes were immersed into liquid nitrogen directly and conserved for at least 24h.After being rapidly thawed in a water bath at 37～40℃for 2min,internodes were washed with sucrose solution(1.2mol/L)for 20min at room temperature.Then the cryopreserved materials were transferred onto 1/2MS+6-BA 0.5mg/·L+NAA 0.1mg/L+GA 0.3 mg/L+sucrose 30g/L+agar 7g/L and cultured in darkness for one week.The heighest survival rate and regeneration rate was 80.00%and 51.11%respectively.3.The suitable resuming medium for hokkaido specialty little fruit tree internodes after cryopreservation was selected.Solid 1/2MS medium supplemented with 6-BA 0.5mg/·L+NAA 0.1mg/L+GA 0.3 mg/L+sucrose 30g/L+agar 7g/L is optimum for the growth of internodes of little fruit tree after cryopreservation.In that condition,survival rate and regeneration rate are the highest,73.33%and 51.11%respectively.