Characterization of the Ipomoelin promoter in sweet potato

Ipomoelin (IPO), a wounding-inducible defense protein from sweet potato, (Ipomoea batatas cv. Tainong 57) can be induced by wounding , Methyl Jasmonate (MJ) and ethylene, and is an excellent target to study plants defense system. The IPO promoter was isolated by iPCR and tail PCR from sweet potato, and contains 1482 bp of the upstream region from the translational start codon of IPO. Five different deletions in the IPO promoter region were created in order to locate its MJ and ethylene-responsive regions. Theβ-glucuronidase (GUS) reporter gene was fused to these promoter segments in the sense direction on PBI221SI100 vector, and the transient expression of these constructs induced by MJ and ethylene in tobacco mesophyll protoplast was studied. Results indicated that the MJ-respond region is from the -552 to -921 bp and the ethylene-respond region is between the -1176 and -1482 bp form the start codon of IPO protein. Also, the regulatory elements of MJ and ethylene in IPO promoter are different.The further 3 new expression constructs each with deletion of one element from the -552 to -921 bp were used to locate the MJ-respond element, the result revealing that the CACACGTC element is one of the responsive element for MJ. The similar result was also confirmed by gel mobility shift assay which point out the involvement of CACACGTC element to MJ response. Besides, the result of gel shift experiment demonstrated the main MJ-responding element is ‘‘TCTAATTTTA,’’ which is similar to the MJ-responding element from Arabidopsis thaliana .