In silico analysis and transient expression of wound-inducible promoter MPI in tomato (lycopersicon esculentum Mill. cv. CH)

Abstract

Various genetic promoters are applied to generate transgenic plants. Constitutive promoters such as CaMV35S are mostly being used; however, the constitutive expression of a gene could be harmful and lead to many changes in plant metabolism, growth and development. Subsequently, using inducible promoters in plant transformation has become more useful. In this project, efficiency of the wound-inducible promoter of Maize Proteinase Inhibitor (MPI) in tomato Lycopersicon esculentum Mill. cv. CH, as a new genetic background, was investigated. Therefore, the MPI was amplified following insertion into pTZ57R/T, sequencing and subsequently ligation into binary vector pBI121. Recombinant plasmids were transferred into Agrobacterium tumefaciens AGL1 strain. The results of in silico sequence analysis of MPI specified methyl jasmonate (MeJA) and abscisic acid (ABA) as responsive motifs, which might play a role in wound signaling pathway. Therefore, tomato leaves were transformed by PBI:MPI:GUS using agroinfilteration approach following GUS expression analysis by applying histochemical GUS and qRT-PCR assays. Our results demonstrated that induction of the MPI promoter only occurred in the wounded leaves and not detected in the control leaves. GUS transcripts in the wounded leaves containing MPI promoter were 4.1 folds higher than those containing CaMV35S, and GUS histochemical assays also revealed that the MPI give higher color intensity (2.64 folds) compared to the CaMV35S. Thus, the MPI could be considered as an acceptable working and inducible promoter to generate dicot transgenic plants harboring insecticidal genes such as different types of cry genes, which just expressed when plants are being attacked by pests.