Molecular Cloning And Expression Analysis Of Gene From Banana Peels Of Differentially Expressed Proteins During Ripening

Banana is one of the largest fruits in southern sub-tropical regions of China. It is atypical Respirationclimacteric fruit.Prolong the storage period and shelf life of banan a, reduce storageand transportation costs, and improve itscommodity Value have been research hotspots in the post-harvest physiology of banana. The research performed a differential protein analysis comparing the total protein extracted from the peels of bananas with and without ethylene-induced ripening treatment in their initial mature stage, and conducted differential protein cloning and expression analysis with molecular biological technology. The conclusion can be drawn as below.1、The total protein of the peels of bananas with and without ethylene-induced ripening treatment in their initial mature stage has been extracted using phenol extraction method. Two-dimensional electrophoresis has been conducted using isoelectric focusing between pH4to pH7. Most of the protein spots we got have oval appearance, and the isolations between them are moderate. The diversity protein spots which exist in bananas pericarp induced by ethylene have been selected by ImageMaster software. Mass spectrum analysis indicated that we have screened the candidate protein which may be relevant to bananas’post-harvest. The candidate protein was denominated as MaCHⅢ.2、According to the peptide information in mass spectrometric data and findings of nucleonic acid sequences relevant to this protein on NCBI, we designed a5’and3’ PCR primer and used bananas total DNA as a template to conduct PCR amplification. We got a1000bp or so amplification product. Sequence analysis shows that the product has a full reading frame of972bp encoding323amino acids residues. This gene sequence does not contain any introns and has99%homology with class Ⅲ acidic chitinase gene. It is certain that MaCHIII gene belongs to class Ⅲ acidic chitinase gene.3、Using qRT-PCR technology to analysis the expression amount of MaCHⅢ g-ene in each tissue and organ of the banana, the results showsit is inconsistent that the expression levels of MachⅢ gene in different tissues and organs of banana. In roots, corm and leaves is only a small amount of expression.Expression levels in flowers, is relatively higher. The expression level is The highest in the fruit. These all illustrates hat MaCH III gene expression istissue-specific. It laid a foundation for the further st- udy of the functions of the gene in fruit ripening and senescence of banana.