The Regulation Of MiR156 On Growth And Development Of Tomato And Lettuce And Its Response To Drought Stress

miRNA is an important member of the gene regulatory network.The mature miRNA consists of only 20-24 nucleotides.It is complementary to target mRNA and then cleaves or inhibits its translation,resulting in the loss of function of target gene.miR156 is a very conserved miRNA in plants.The most studied target gene of miR156 is the SBP transcription factor,which plays an important role in the growth and development of plants.However,the main work at present is concentrated on the model plant Arabidopsis,and there is little related work in crops.In this research,the effect of tomato explants on the transformation efficiency of miR156 was studied and some transgenic materials were obtained.It was explored whether the target Sl SBP5 transcription factor of tomato miR156 regulates the expression of miR172.In addition,differences in phenotypes and drought tolerance between overexpressing MIR156 and target mimic STTM156 transgenic lettuce lines were compared.This study lays the foundation for further exploration of the function of miR156 in crop growth and development and stress response.The main findings are as follows:1.Four vectors for overexpressing OX-MIR156 a,OX-MIR172 a,target mimic STTM156 a and STTM172 a were successfully constructed.The genetically modified tomato “Moneymaker” plants were obtained : 8 lines of OX-MIR156 a,2 lines of STTM156 a,6 lines of OX-MIR172 a,and 4 lines of STTM172 a.Genetically modified tomato “Micro-Tom” plants were also obtained: 20 lines of OX-MIR156 a,8 lines of STTM-156 a,10 lines of OX-MIR172 a,and 5 lines of STTM172 a.2.Different transformation systems and Agrobacterium containing different genes were used to infect tomato explants to optimize the genetic transformation system.Overexpression of MIR156 a and STTM156 a showed different differentiation efficiencies in different systems.Overexpression of MIR156 a and target mimic STTM156 a showed different differentiation efficiencies in different systems.After infestation with OX-MIR156 a,ZT(1 mg/L)+ IAA(0.1 mg/L)induced cultured cotyledons had high differentiation efficiency,while explants infected with STTM156 a had higher differentiation efficiency when ZT(2 mg/L)+ IAA(0.2 mg/L)induced hypocotyls.Taken together,the OX-MIR156 a system had a higher efficiency of induction differentiation.Different tissues of miR156 may affect the differentiation efficiency of different tissues through the response to plant hormones.3.The yeast single-hybrid technique was used to verify whether the transcription factor Sl SBP15 binds to the promoter region of sly-miR172.The results showed that there was no activation in this study,and the promoter might not be involved in the expression regulation of downstream genes.4.Phenotypic observation of transgenic lettuce showed that the expression level of miR156 affected seed development,leaf morphology,and anthocyanin and chlorophyll levels.OX-MIR156 line had more obvious phenotypic differences than wild type strains.The seeds were larger than the wild type;the leaf initiation rate was fast;the leaves were small and slender;the number of trichomes was increased abnormally on both abaxial and paraxial surfaces;the anthocyanin and chlorophyll levels were increased.miR156 inhibited the expression of Ls SPL.It is speculated that the change of Ls SPL expression may affect the expression of a variety of downstream genes that respond to plant growth and development,thus showing different phenotypes.5.Plants of wild type,STTM156-1 and STTM156-4 lines of lettuce were subjected to natural drought.The results showed that after 15 days of drought,the expression of miR156 was increased and the expression of Ls SPL9 was decreased in wild type lettuce.The leaves of different genotypes became smaller under drought,and the old leaves turned yellow and dry.After drought treatment,the contents of MDA and hydrogen peroxide in the leaves of two STTM156 strains increased significantly,the activity of SOD decreased,and the activity of POD increased lower than that of wild type,indicating that the target mimic STTM156 strain was more sensitive to drought stress.