| MircoRNAs are a class of small RNAs of about 21-23 nucleotides in length that are not encoded.It can base on the mRNA base pairing with its corresponding target gene to guide the degradation of its mRNA silencing complex(RISC)to hinder translation,thereby regulating the expression of its corresponding gene at the post-transcriptional level,miRNA regulation in plant development,adversity Responses and hormone regulation play an important role.Potassium is one of the essential nutrients for plant growth and development.Potassium deficiency not only limits plant growth but also reduces yield.By analyzing the expression profile of small RNA in two tomato lines with different potassium utilization efficiency,it was found that there was a significant difference in the expression of miR168 a in the two strains.Combined with previous studies,we speculate that the miRNA may play an important role in the regulation of tomato under low potassium stress,but there is no related report on the regulation of miR168 on low potassium stress.In order to elucidate the biological function of tomato miR168 a,the tissue specific expression patterns of mi R168 a and its target gene AGO1 in potash sensitive and low potassium tolerant tomato are detected by qRT-PCR.JZ18 of potassium sensitive tomato is used as the material,and the Sly-pre-miR168 a sequence is obtained.Gene specific primers are designed to amplify Sly-pre-miR168 a.The obtained PCR fragment is digested by SmaI and Bam HI and link to the overexpression vector pBI121.Agrobacterium tumefaciens mediated leaf disk transformation of tomato 35S::Sly-miR168 a overexpression T0 plants are obtained and identified by potassium free water culture.The main results are as follows:(1)Tomato JZ34 have no significant changes in tomato under low potassium stress and could grow normally.JZ18 have a significant decrease in stem length,root length,fresh matter weight and dry matter weight under low potassium stress.The content of potassium ion accumulated in JZ18 tomato is lower than that in JZ34 tomato.The potassium absorption rate of JZ18 is higher than that of JZ18 in potassium depletion test.The results of tomato seedling indicates that there are genotypic differences in the ability of tomato to potassium in seedling stage.It reflects the sensitivity of tomato plants responding to potassium nutrition.(2)The expression level of miR168 a in tomato JZ18 decreased significantly after potassium-free treatment,and decreased by 9.4 fold at 7 days.On the other hand,the expression level of JZ34 tomato increased without potassium treatment,which increased by3.8 times at 7 days,indicating that miR168 a had a certain effect on improving tomato tolerance to hypokalemia.From the trend of gene expression,the expression of AGO1 is exactly the same as that of miR168 a.(3)Compared with wild type tomato plants,the expression of miR168 a in transgenic plants was significantly increased,while the AGO1 expression level of mi R168 a target gene predicted by bioinformatics is decreased accordingly.Overexpression of transgenic tomato of miR168 a and wild tomato are used as materials.After 7 days of potassium deficiency culture,the lateral roots,root hairs and surface area of transgenic lines are increased and no chlorotic phenotype appeared at the leaf edge.The results indicates that the ability of low potassium tolerance of transgenic lines is enhanced. |
