| Banana (Musa paradisiaca L.), one of the four widely grown crops world, which has the higher economic value, is the tropics agricultural mainstay industry. The banana land of our South China had received destructive damage because of the cold wave invasion, which causing severe losses in economic production. The one of main breeding goals of banana is to cultivate cold resistant variety. But at present the newly cultivated cold resistant variety has not radically solved banana's cold resistant problem. Therefore, the banana cold resistant mechanism research is urgently need strengthen in order to provides the theory basis of cultivating the new cold resistant banana variety, reducing the losses in economic production and expanding plant areas.The experimental material of our research is used the main cultivate variety Musa AAA Cavendish cv baxi. The banana seeding had firstly been pretreated via the different density ABA and BR and then decreased the temperature to 7℃and maintained 12 hours. The conductivity, soluble sugar content, soluble protein content, MDA content, SOD activity and chlorophyll content of the banana seedling were mensurated and the banana seedling shape, the functional foliage shape as well as the cellulary tissue dissection structure of leaf were investigated. At the same time, the difference fragment between treatment and control were cloned and these cloned fragment were compared with the sequences of nucleic acid and protein database in NCBI and obtained some cold-correlation clone fragment. Using 3'-RACE and 5'-RACE technology, the rbcS-mc gene were cloned and the biological information study, RT-PCR and transfers the gene analysis of the gene obtained were analysis. The the main results were as follows:1. The banana seedling were treated with the different ABA density such as 5 mg·L-1,10 mg·L-1,15 mg·L-1,20 m g L-1·,25 mg·L-1mg and different BR density such as 0.3mg·L-1,0.6 mg·L-1,0.9 mg·L-1,1.2 mg·L-1,1.5 mg·L-1 under the cold-stressed condition. Compared with the control, the treatment in certain treatment density may obvious reduce the electrolyte leakage rate, increase SOD activity, enhance the soluble protein content at a certain extent, slow down the MDA content change, distinct enhance soluble sugar content and delay the chlorophyll degraded speed of the banana seedling leaf blade. These metabolite change have played extremely key and the important function to enhances banana seedling cold-stressed period resistance. The best protected effect density respectively is 20 mg·L-1 ABA and 0.9 mg·L-1 BR.2,The plant exterior configuration exhibit obvious difference after 12h under 7℃cultivated. The tested plant growth vigorously and healthily, the leaf color present shine green, cold impair degree is 0 levels, cold impair index is 4.5% under the 20 mg·L-1 ABA spray (treatment 4) and the treatment effect is the best; while the control plant have 75% leaf blades completely wither, the other leaf blades have the slight frostbite, cold impair degree is 3 levels, cold impair index is 76% and the difference is remarkable. The tested plant growth well, cold impair degree is 0 levels, cold impair index is 4.1% under the 0.9 mg·L-1BR spray and the treatment effect is close to treatment A4. The result show that the employment of a certain density BR or ABA may enhance cold-resistance in the certain degree of the banana seedling, but enhances effect do not presents direct ratio with ABA or the BR. In the dissection structure aspect, along with the continual cold-stressed, banana leaf blade cellular tissue structure tightness presents a more orderly change, leaf blade thickness has changed attenuation. The leaf blade's protection structure such as cuticle tissue, stockade tissue present incrassated tendency under the certain degree cold-stressed with BR treatment compared with control, and the CTR value also reduces from 51.1% CK to the 47.5% BR.3. Using the ADGE technology, the 18 special expression DNA fragments in the cold-stressed banana seedling under the BR induction were cloned and sequenced. Clones 2-1T has shared 99% consistency in the 466bp nucleotide sequence of ribulose-1),5-bisphosphate carboxylase/oxygenase small subunit (rbcS1) No AF008214 in NCBI when search in the homology at three big international nucleic acid databases, The banana ribulose-1,5-bisphosphate carboxylase/oxygenase small subunit gene (rbcS-mc) was obtained using TaKaRa 3'- RACE and 5'- RACE reagent Separately. With DNAstar software, the sequence as well as the known clone piece sequence spliced to contain the complete code area of ribulose-1,5-bisphosphate carboxylase/oxygenase small subunit gene (rbcS-mc) sequence and some biology information analyses were carried on. Meanwhile, the RT-PCR expression analysis of the banana rbcS-mc gene in the banana plant different situation has carried on. The RT-PCR analysis result showed this gene in the banana plant's leaf blade's expression quantity is higher than leaf sheath, while in the root does not express the gene, which indicated the banana rbcS-mc gene has the organization specificity expression in the banana plant.4. According to banana rbcS-mc gene with integrity ORF sequence, the primers have been designed. Using the banana rbcS-mc gene DNA as the template, the banana rbcS-mc gene with integrity ORF sequence has been cloned and constructed included the integrity code area plant to expressing vector, then transformed agroacterium GV3101 and transformed tobacco, finally gained the transgenic tobacco plant. These transgenic tobacco plant will help to further study of the banana rbcS-mc gene character and function.
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