| The fruit specific promoter play a crucial role that improve the fruit’s quality traits in turning genetechnology. The separation of citrus fruit ripening-specific promoter and analysis of its function has greatsignificance which can reveal the fruit mature molecμlar mechanisms and use the biotechnology to improvefruit quality. But Citrus fruit ripening-specific promoter is not reported. On the basis of getting citrus fruitripening-specific genes in the early, we separated the5’ upstream promoter region from the gene withGenome Walking technology, and forecasted its functional components by bio informatics software; webuilded three different promoter regionals to drive intron-GUS, we preliminary analyzed the expression andregμlation of the different promoter regions in various tissues and organs of citrus by the transientexpression system. It provides the basis for further ianalysis and use of promoter function. The main resμltsare as follows:(1)We separated and obtained the citrus CsPMEI-InvI gene5’ upstream promoter sequence by GenomeWalking. The analysis of bio informatics showed that the promoter gene is1159bp, and containing13CAAT-box,18TATA-box,11acting elements about lighting reaction, cis-acting elements induced byanaerobically, enhancer elements which inducting by hypoxia-specific,cis-acting elements which regμlatingby biological clock, the myoglobin binding sites in drought reaction, the cis-acting elements oflow-temperature reaction, and the cis-acting elements which with JA reaction, the cis-acting elements thatexpress by endosperm, cis-acting elements that react by salicylic acid and8functional elements of unknownfunction, totally30functional elements with different types. So we inferred that CsPMEI-InvI genepromoter sequences have the basic structure and function of promoter, is the promoter region of maturecitrus fruit’s specific genes CsPMEI-InvI.(2) To build the plant expression that suitable for transient expression, we chose pCambia1301whichcontaining iGUS gene as based vector. IGUS gene is not expressed in agrobacterium, and it can avoid theinterference caused on the study of plant transient expression.According to the5’ upstream region,wedesigned specific primers, and cloned three different promoter region fragments, and replaced them withCaMV35S constitutive promoter, then we constructed there expression vector pLZ25, pLZ26, pLZ27.(3) We established GUS staining transient expression system by agrobacterium-mediated way,preliminary analysis of the regional role of different promoters in the kumquat tissues and organs.According to the staining resμlts, initially concluded that the promoter region has strong fruit’s expressioncharacteristics, but it needs further verification by transgenic. |
