| Banana has been ranked as the world's fourth most important food crop and has very important economic value. Besides, it is an ideal natural bioreactor. Banana is being engineered to produce antigens so that they can be used as oral vaccine . But it is a difficult challenge for traditional plant breeding because bananas produce fruit without pollination and reproduce vegetatively, and almost all important banana cultivars have three sets of chromosomes, making it difficult to add desirable traits by classical breeding. So, it is very important to develop transgenic technology in banana. We aimed in this study to isolate and identify the effective promoter which is adaptive to banana transgenic research to develop the transgenic technology in banana.We isolated a banana actin gene fragment named actinl from genomic DNA by the method of PCR. DNA sequence analysis and homology comparison .indicated that it has a 99% homology with the actin gene which has been submitted to GenBank.After this, an 5' upstream promoter region of actinl gene was isolated by polymerase chain reaction from genomic DNA . Both the two fragments of the promoter were cloned and sequenced. we analysed the core promoter region and the upstream regulatory elements in the fragments with the software of PROMOTER PREDICTION and PLANT CARE .By fusing this two fragment with gus reporter gene and nopaline terminator, we get two transient expression vectors which named pCAMBIA-ACTP and pBI-ACTPS.The two constructs and PBI121 were delivered into the tissue ( root, leaf,fruit ) of banana for transient expression by particle bombardment . Transient expression studies in transgenic banana showed that both the actinl promoter fragments can drive strong reporter gene expression in root, leaf and fruit of banana. Also, the colorimetric assay had been carried out to make further research on the difference of the two fragments in their promoting activity, and the results supported the conclusion drawn above. |
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