| To date,the research of generation and genetic transformation of bamboo were focused on woody bamboos.However,woody bamboos had long long-periodic,complicated flowering phenomena,low seed setting percentage and low efficient of regeneration,which dramatically inhindered the genetic transformation and veritifiacation of function of bamboo genes.In this study,after the biological characteristics of introduced herbaceous bamboo,Mniochloa abersend,was observed,the regeneration system was estabilished by by using shoots as explants,which were easily to get.In addition,various basal media and plant growth regulators were tested for the purpose of selecting out the optimistic culture conditions for callus induction and proliferation,shoot differentiation,root induction and getting regenerated plantlets of M.abersend.On the basis of this stable and efficient system,some genetic transformation indicators were teated to establish the genetic transformation system of M.abersend,providing a new approach for verification of function of bamboo genes.The main results are as follows:1.Biological characteristics of M.abersendM.abersend was perennial and tufted herbaceous bamboo,which grows not so well under 0? environment.Besides,M.abersend blossomed and set seeds annually,which had spike-shaped raceme,belonging to the genuine inflorescence,whose basic units were twin spikes.2.Regeneration system of M.abersend?1?The optimal medium of callus induction was MS basal medium supplemented with 1 mg·L-1 2,4-D,1 mg·L-1 NAA,30 g·L-1 sucrose and 8 g·L-1 Type A agar.The frequencies of callus and compact callus induction were 86.67% and 63.3% respectively.The calli obtained grew well and had high capacity of proliferation.?2?The optimal medium for callus proliferation was MS basal medium supplemented with 0.1 mg·L-1 2,4-D,30 g·L-1 sucrose and 8 g·L-1 Type A agar.The proliferation times of callus was 8.01 with generating a host of embryoids.?3?M.abersend showed 100% differentiation rate.The optimal medium for callus differentiation was MS basal medium supplemented with 1 mg·L-1 KT,1 mg·L-1 NAA,30 g·L-1 sucrose and 3 g·L-1 gelrite,in which the shoots were green and strong.?3?M.abersend showed 100% rooting rate.The optimal rooting medium of regenerated plantlets was 1/2 MS supplemented with 0.5 mg·L-1 IBA,30 g·L-1 sucrose and 3 g·L-1 gelrite.Roots were generated 3 days after inoculated.Moreover,the roots were long and thick,which were easily to be transplanted.?4?The survival rate of hardening regenerated plantlets was 90% after transferred to an equal ratio mixture of peat,vermiculite and perlite,with the plants growing vigorously.3.Genetic transformation of M.abersend?1?The optimal condition of genetic transformationAfter 3-days-preculture in liquid medium,the compact calli were infected 20 minutes in the Agrobacterium tumefaciens LBA4404,whose OD600 was 0.5.The callus were selected after 3-days-coculture for a month.Finally,the resistant callus rate was 34.67% and the callus grew well.?2?The resistant calli were devided,some were used to be test by PCR and the rest were prepared to differentiation.The conversion rate of resistant calli was 26.67%,which proved the gene cod A was transformed into the callus of M.abersend.Further test would be taken when the resistant calli were differentiate to plantlets. |
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