Regeneration System Via Shoot-tips Of Dendrocalamus Asper And Genetic Transformation

In this study,the optimum medium and culture conditions of callus induction,differentiation and root induction of regenerated seedlings was studied with the shoot tips of Dendrocalamus asper,and the efficient and stable regeneration system was established.According to the regeneration system,the genetic transformation conditions of D.asper were explored,and laying the foundation for genetic improvement of bamboo.The main results are as following:1.Regeneration system of callus induction in shoot-tips of D.asper（1）The suitable medium for callus induction in shoot-tips was MS medium supplemented with 3 mg·L^-12,4-D and（0.5-1）mg·L^-1NAA,and the induction rate of granular and compact callus was 61.7%.（2）Low light(10μmol·m^-2·s^-1)treatment had a greater effect on callus differentiation,and the appropriate treatment time was 6 days.And the suitable shoot differentiation medium was obtained in MS medium supplemented with 1 mg·L^-16-BA,1 mg·L^-1KT,and 1 mg·L^-1NAA with the highest differentiation rate of 46.67%,and the regenerated shoots grew well.（3）The appropriate rooting medium was 1/2 MS medium supplemented with 3 mg·L^-1IBA.The rooting rate was 55%,and the roots were thick and some of them have lateral roots.And the rooted plantlets were transferred to artificial mixture（vermiculite:perlite:peat=1:1:1）with the shoot survival rate up to83.33%.2.The genetic transformation system D.asper via Agrobacterium tumefaciens The CodA gene was transferred into the receptor,callus of shoot-tips from D.asper by Agrobacterium tumefaciens-mediated method.The results showed that after 20 mg·L^-1,40 mg·L^-1hygromycin concentration ladder screening,the rate of resistance callus was as high as 51.9%while co-cultivation for 3 days.And some resistance plants were obtained,one of the regenerated albino plants were proved positive through PCR.The regeneration of green plants was too small to be verified.The maize Leaf color（Lc）gene was transferred into the receptor,callus of shoot-tips from D.asper by Agrobacterium tumefaciens-mediated method.The callus of D.asper shoot-tips was selected by hygromycin and transferred to differentiation culture.A purple regenerated plant and a purple resistant callus were successfully obtained.3.Preliminary study on the genetic transformation system of D.asper mediated by gene gun The gene-gun-mediated method was used for the first time in the establishment of the genetic transformation system of bamboo,and the green fluorescent protein gene was introduced into the callus of shoot-tips from D.asper.The instantaneous fluorescence of green fluorescent spot could be observed after 3 days of culture.