SlmiR168a Mediated SlAGO1 Regulates The Molecular Mechanism Of Tomatoes Low Potassium Stress

Tomato(Solanaceae Lycopersicon esculentum Mill)is inseparable from the supply of potassium during its development stage.Potassium deficiency will seriously affect the quality and quantity of its fruit.But nowadays,there is a serious shortage of potassium elements that can be absorbed and used in farmland soils in China.Therefore,cultivating tomato varieties with low potassium tolerance is an effective method to improve the potassium absorption efficiency of tomato and adapt to potassium-deficient soils.The research team obtained low-potassium-sensitive tomato JZ18 and low-potassium-tolerant tomato JZ34 by identifying the biomass and yield of tomato under low-potassium stress.Through analysis of omics and transgenic tomato materials,it was found that miR168 a affected the lateral roots of tomato under low-potassium stress.Root hair development,improve low potassium tolerance.Studies have shown that SlAGO1 is a target gene of tomato miR168 a.However,the molecular mechanism of miR168a-regulated AGO1 in tomato resistance to low potassium stress is unknown.In this study,35S: mSlAGO1(mutant AGO1)transgenic tomato material was constructed by site-directed mutation of SlAGO1 so that it was not regulated by miR168 a and did not change its amino acid sequence.Transgenic lines and miR168 a overexpression homozygous lines were identified for potassium content,chlorophyll determination,and leaf and root hair development.Furthermore,35S: SlmiR168 a and 35S: mSlAGO1 were used for miRNA sequencing to analyze the pathway of miR168a-mediated SlAGO1 to regulate downstream miRNA induced by low potassium stress and its target genes.The research results provide new ideas for in-depth analysis of the molecular mechanism of tomato tolerance to low potassium,as well as new strategies for the selection and breeding of new tomato varieties with low potassium tolerance.The main findings are as follows:1.RT-PCR detection found that AGO1 was expressed in both the rhizomes,leaves and fruits of tomato.It was the highest expressed in leaves and the least expressed in fruits.During the fruit development process,the expression level from high to low is the color change period> green ripe period> red ripe period.2.Construction of SlAGO1-GFP vector,transient transformation of tobacco leaves for subcellular localization,the results showed that SlAGO1 localized in the cytoplasm and nucleus.3.Construct 35S: mSlAGO1 transgenic tomato lines and test the T0 generation to obtain 4 positive lines.4.After low potassium treatment,35S: SlmiR168 a showed more root hair,higher root-shoot ratio andhigher chlorophyll content to enhance tolerance to low potassium stress;while 35S: mSlAGO1 had less root hair and root-shoot ratio Decreased,chlorophyll content decreased,showing more sensitive to low potassium stress.The determination of potassium content showed that 35S: SlmiR168 a was significantly higher than that of the control group JZ18,while the potassium content of the 35S: mSlAGO1 transgenic line was significantly decreased with the lowest content.5.miRNA sequencing of the above 35S: SlmiR168 a,35S: mSlAGO1 transgenic materials,and the results showed that differential miRNAs were significantly enriched in DNA-templated transcriptional regulation,protein phosphorylation,redox processes,abscisic acid response,cell differentiation,and growth Reaction,metal ion binding,etc.Among them,the known Significantly different expressions miRNAs include: miR393 is specifically expressed in 35S: mSlAGO1,and miR156 is significantly up-regulated in 35S: SlmiR168 a.It was significantly down-regulated in 35S: mSlAGO1.This result indicates that miR168a-mediated SlAGO1 may affect downstream miR156 and miR393 regulatory pathways in response to low potassium stress.