Establishment Of Technique System Of Agrobacterium-mediated Transformation Cotton (Gossypium Barbadense L.) And Identification Of Resistance To Cotton Boll Worm Of Bt Transgenic Cotton

In this study, we studied the difference on regeneration by somatic of land cotton(simian3,kezimian312) and island cotton (xinhai30,xinhail6) and finded the difference of culture prosess on the two cultivars, We maked hereditary transform of Bacillus thuringiensis toxin protein gene (Bt) by Agrobacterium.In order to conform foreign gene have integrate into genome by PCR on T1 and RT-PCR on T2. We reguired transformatic plantet and Identfied the resistance worm to requiring transformationg planet. The results of worm feeding intial certificate this.The results of this research such as:1.We used hycopotys of island cotton and land cotton as explant.We used Screening the best condition of embryo callus induction condition under the same methed of callus induction and proliferation. The best embryo callus induction mediun on land cotton is MSB adding 1.9 g/L KNO3, The rate of embryo callus induction reach 70%. The best embryo callus mediun on island cotton is MSB adding 1.9 g/L KNO3 and 0.3mg/L NAA. The best embryo callus multiplication mediun on land cotton is MSB adding 1.9 g/L KNO3, 0.1 mg/L Glu and 0.1mg/L Asn. The best embryo callus multiplication mediun on island cotton is MSB adding 3.8 g/L KNO3,0.1 mg/L Glu and 0.1 mg/L Asn.2. The best embryo Somatic embry differentiation mediun on island cotton is MSB adding 1.9 g/L KNO3, 0.1 mg/L Glu,0.1 mg/L Asn and 0.05m/L IBA.3. Arobacterium-mediated embryo callus of island cotton xinhai30 and xinhai 16, Requiring BT gene transgenic plants. Converting conditions for:Island cotton embryo callus was soaked bacterium fluid with OD of the value of 0.5～0.6,15min. post-communist 24h. Resistent embryo callus will turn into adding 50mg/L kana and 400mg/L cephalosporins amphotericin medium induced somatic embryos genetically differentiation. The best seedlings medium of resistance cotyledons embryo is 1/2MSB adding 300mg mg/ L cephalosporins amphotericin.4. We screened the kanamycin resistence on the transgenic plants of T1 generation.The results shows that resistance rate is 12.6%. Detection by PCR detection on T1 transgenic plants of positive kanamycin resistence Then identify the insect resistance of T1 generation transgenic plants, insect resistance results show that transgenic island have insect resistance. We got T2 generation the transgenic plants tissue seedlings by tissue culture of T1 generation of transgenic island cotton. Identify T2 generation of the transgenic plants by RT-PCR detection, the results showed that Bt could be normal expression in transgenic T2 plants.