Study Of Tissue Culture Of Hippeastrum ‘shengyin NO.3’ And Globba Winitti And Betula Halophile

In recent years,due to the rapidly developing of flower market,it needs more excellent new species and varieties,and plant species with extremely small populations also need to solve the problem of propagation.Using tissue culture technology can produce lots of seedlings to meet the market demand and help to realize the utilization of plant resources.In this experiment,Hippeastrum‘Shengyin No.3’,Globba winitti and Betula halophila were used as materials to establish rapid propagation system in vitro so as to provide technical supports for large-scale production.The main results are concluded as follow:（1）The bulblet of H.‘Shengyin No.3’was used as explant.Bulblet was initially wiped with 75%（v/v）alcoholic,followed by immersed in 75%（v/v）alcohol for 30 s,and then in0.10%（w/v）HgCl₂ three times for 5 min+3 min+2 min,respectively.Through this method,a higher success rate for explant disinfection could be obtained.The medium for induction of bulblets was MS+5.00 mg/L 6-BA.The LS is the most suitable basic medium for proliferation and growth.The bulblets cut into 8 parts could be induced into the most little bulblets,the proliferation rate were 14.28 and 15.11 for 30 d and 60 d culture respectively.The most effective plant growth regulator for bulblet proliferation was 6-BA.The bulblet multiplication ratio was 11.07 after 60 days of culture which were divide into 4 parts in LS+4.00 mg/L 6-BA.The seedlings obtained highest fresh weight incrementon in the MS+4.00mg/L 6-BA+1.50 mg/L NAA.H.‘Shenyin No.3’cultured in MS+2.00 mg/L PP₃₃₃ had short,wide leaves and large bulbs,which played a good role in inhibiting leaf growth and promoting bulb expansion.In MS+4.00 mg/L 6-BA medium.The maximum bulb diameter was 7.94 mm in MS+4.00 mg/L 6-BA supplemented with 120 g/L sucrose.The optimal rooting medium was MS+2.00 mg/L IBA,the rooting rate was 100%.H.‘Shengyin NO.3’was easy to survive in transplanting,and the survival rate was 100%in the 14 substrates tested,and H.‘Shengyin NO.3’growed well in the matrix of peat soil,coco peat:pearl:vermiculite with volume ratio 1:1:1 and peat:perlite:coco peat with volume ratio 1:1:1（v:v）.（2）Young flower buds and sprouts from rhizomes of G.winitti were used as explants.Explants were initially wiped with 75%（v/v）alcoholic,followed by immersed in 75%（v/v）alcohol for 30 s,and then in 0.10%（w/v）HgCl₂ three times for 5 min+3 min+2min(young flower for 3 min+2 min+1 min,respectively.Through this method,a higher disinfection success rate can be obtained.Explants primary culture with MS+2.00 mg/L 2,4-D+0.50mg/L 6-BA and MS+5.00 mg/L 6-BA+0.01 mg/L NAA,and after 30 days,20.00%and80.95%of adventitious bud induction frequency were obtained respectively;The adventitious buds induced by the two explants had no significant difference in the multiplication ratio.The most suitable proliferation medium was MS,and the adventitious buds proliferation ratio at 0.10 mg/L TDZ was16.46 by direct somatic embryogenesis.The suitable inoculation density were 15 single buds per bottle.After 30 days of culture,the proliferation ratio of adventitious buds was 7.28.The best rooting medium was 1/2 MS+0.50 mg/L NAA,the rooting rate reached 100%.The best transplanting substrate was peat soil:vermiculite=2:1（v:v）,the survival rate was 100%.（3）The stem tips and stem segments of B.halophila were used as explants.Explants were initially wiped with 75%（v/v）alcoholic,followed by immersed in 75%（v/v）alcohol for 30 s,and then in 0.1%（w/v）HgCl₂ three times for 5 min+3 min+2 min,respectively.Through this method,the minimum contamination rate was 34.00%.The most suitable medium for adventitious bud induction of B.halophila was B₅ medium;the best inoculation density per bottle in subculture medium MS+0.50 mg/L 6-BA+0.05 mg/L NAA was 20-25 stem segments with two nodes,but the adventitious buds were thin.The optimal proliferation medium was MS+0.50 mg/L 6-BA with a proliferation ratio of 31.04.Strong seedlings of B.halophila were favored in medium with 1/2 MS and MS,supplemented with0.50 mg/L IBA.The most suitable sucrose concentration was 30 g/L and 45 g/L on strengthing seedling culcure.The roots are thick and strong on the MS medium with 0.50mg/L PP₃₃₃.The light intensity of 3000 Lux was the best for strengthening seedling culture of B.halophila;the best rooting medium was MS+0.30 mg/L NAA,the rooting rate reached100%.The best transplanting substrate was peat soil:sand=1:1（v:v）.the survival rate was 95%and the plantlets were robust.