The Interaction Mechanism Of MiRNA And Its Target Genes Under The Stress Of Marssonina Populi

Poplar is a kind of important woody tree, which has great economic andenvironmental value. The poplar black spot disease, caused by the fungusMarssonina (Marssonina brunnea), is one of the main diseases that occur in genusPopulus. At present, the molecular mechanism of poplar resistance to poplar blackspot disease, especially of miRNAs involved in this regulatory network is poorlyunderstood. The study of the molecular mechanism of poplar black spot disease willprovide theory and practice basis for the prevention and cure. In this study, based onpoplar black spot disease-resistant P. euramericana "Nanlin895", of the miRNAsrelated to poplar black spot disease resistance and their molecular mechanism, themain results were as follows:Combined with disease resistance related miRNAs reported in the literature, wepredicted miRNAs related to resistance to poplar black spot disease, and designedfungus infection experiment, using the stem loop RT-qPCR (stem loop primersreal-time quantitative reverse transcription PCR) assay for the identification. Theexpression level of miR156a and miR164a in Nanlin895was significantly suppressedafter fungus infection. We selected miR164a to clone its precursor MIR164a genes.By the use of RACE technique, we obtained the full-length sequence ofPdNAC1, the predicted target gene of miR164a. Through the qRT-PCR assay to theexpression of PdNAC1, results showed that PdNAC1was marked increase in theamount of expression after fungus infection in Nanlin895. The expression trend ofPdNAC1is negatively correlated with miR164a. Using the transient expressionsystem of poplar protoplast, we studied the cellular localization of PdNAC1, theresults showed that the PdNAC1transcription factor is mainly located in the cellnucleus, but also expressed in the cell membrane at the same time.Based on the mature poplar protoplast transient expression system, we initiallyestablished a simple and intuitive target identification method, and used for thetarget identification of PdNAC1. According to the results, through the multiplemutations on the target locus of PdNAC1, can effectively remove the expressionsuppression in PdNAC1by miR164a, verified the prediction of target locus onPdNAC1.