Micropropagation Of Stem Tips And In Vitro Regeneration Of Leaves In Red-Flowered Strawberry

The plants of the genus Fragaria bloom white flowers.The red-flowered strawberry is the intergeneric hybrid from the cross of Fragaria and Potentilla with good ornamental value and development potential.Their flowers are rich in colors from pink to red,and the fruits taste delicious.The red-flowered strawberry could be used as ground cover or pot flower now.The stem tips of stolons in red-flowered strawberry cultivar ‘Sijihong’ were used as explants in this experiment.The effects of culture media and exogenous hormones on the primary culture,multiplication and rooting culture were analyzed,and the in vitro rapid propagation system was established,which laid a foundation for the breeding of red-flowered strawberry.It was of great significance to realize the industrial propagation of red-flowered strawberry.The leaves of ten red-flowered strawberry cultivars,such as ‘Sijihong’,‘Pink Beauty’,‘Pretty Beauty’,‘Pink Princess’,‘Xiaotaohong’,‘Red Rosa’,‘Meihong’,‘SF14’,‘26SF’,and ‘X1-3’were used as explants.The effects of genotype,plant growth regulator,dark culture,inoculation method and Ag NO3 concentration on callus,differentiation and plant regeneration were analyzed.The results were as follows:The stem tips of red-flowered strawberry cultivar ‘Sijihong’ were established as micro propagation system.The effects of three primary culture media: 1/2MS,MS and MS+BA 0.5mg·L-1 on the germination rate were analyzed.The results showed that the germination rates of MS and MS+BA 0.5 mg·L-1 were both higher than 1/2MS with no difference,64.43% and65.33%,respectively,in which the primary culture media MS+BA0.5 mg·L-1 could produce a small amount of proliferation.The effects of different hormone combinations on the expansion and growth of red-flowered strawberry ’Sijihong’ were compared.The best subculture medium was MS+BA 0.5 mg·L-1 + NAA 0.1 mg·L-1.The proliferation coefficient was 4.23,and the average height reached 3.80 cm;The optimal rooting medium was1/2MS+NAA 0.1 mg·L-1 with rooting rate 98.50%,and the average root number was 18.23;After 5d of domestication,the tissue culture plantlets were inserted into the sand: vermiculite= 1:1.After 28 d,the statistical survival rate was 83.9%,and the plant grew vigorously and the root systems developed well.The leaves of ten red-flowered strawberry cultivars,such as ‘Sijihon’,‘Pink Beauty’,‘Pretty Beauty’,‘Pink Princes’,‘Xiaotaohong’,‘Red Rosa’,‘Meihong’,‘SF14’,‘26SF’ and‘X1-3’ were used as explants to establish in vitro regeneration system.The results showed that the genotype was the main factor affected the regeneration of adventitious shoots of red-flowered strawberry leaves.Among the ten red-flowered strawberry cultivars compared,six cultivars were regenerated adventitious shoots.The adventitious shoot differentiationrate from high to low was ‘Pink Beauty’ > ‘26SF’ > ‘Pretty Beauty’ > ‘14SF’ > ‘Xiaotaohong’> ‘Meihong’.By comparing the effects of different hormone ratios on leaf callus and adventitious shoot induction,the combined effect of TDZ was better.And the combination of TDZ and NAA was better than the IBA combination.The optimal medium for callus induction and adventitious shoot induction of the cultivar ‘Pink Beauty’ was MS+TDZ 1.0mg·L-1+NAA 0.2 mg·L-1.The adventitious shoot differentiation rate could reach 44.57% with an average of 2.93 adventitious buds per leaf regeneration;The leaves were inoculated two types,the reverse surface and the front surface contacted medium,respectively.The results showed that the orientations had no effect on regeneration of leaves.However,when the front surface was in contact with the media,the middle part of the leaf was arched,and the leaf edge wound could contact the medium,which accelerated the induction process.When the culture is started until 14 d,the callus was germination,and at 55 d,adventitious buds began to occur.When inoculated on the reverse side of the leaf,the leaf surface was U-shaped,the leaf edge could not contact the media,and the induction speed was relatively slow;The leaf induction of adventitious shoots was compared under different dark culture days.It was found that dark cultured for 14 d could effectively increased the rate of adventitious shoot differentiation.The adventitious shoot differentiation rate was 63.67% and the average number of adventitious shoot was 3.27.When the dark treatment days were more than 14,the adventitious shoot differentiation rate showed a downward trend.The dark treatment was significantly lower than the control group at 10.12%,when the dark treatment days were 28.By adding a certain concentration of Ag NO3 into the media,when the Ag NO3 was 2 mg·L-1,the regeneration frequency of the leaves could be increased,but as the concentration continued to increase,the regeneration of adventitious shoots was inhibited.