Functional Verification Of Cadmium Responsive Histone Demethylase Gene SlJMJ2 And SlJMJ12 In Tomato

Cadmium(Cd)is a heavy metal that is an unnecessary element in the growth and development of plants.Excessive accumulation of cadmium in soil can affect plants and even cause death.Cadmium accumulation in plants can eventually enter the human body through the food chain,endangering human life and health.Tomato(Lycopersicon esculentum Mill.)is an annual herb of the genus Tomato in the Solanaceae family.It is a widely cultivated fruit and vegetable plant of important economic value worldwide,with high nutritional value,and has the largest planting area in China worldwide.However,in recent years,due to the excessive application of fertilizers and pesticides,the problem of heavy metal residues in cultivated soil has become increasingly prominent,which not only affects the growth and development of plants but also affects human life and health.Histone demethylases(JHDM)are a family of proteins that contain only the Jmj C domain.However,at present,there are more studies on the response of histone demethylases genes to salt and alkali stress,and fewer studies on heavy metal stress.Therefore,it is important to study the role of histone demethylase in tomato resistance to Cd stress.The results of this test indicate that:(1)Cloning and obtaining the full length CDS of SlJMJ2 and SlJMJ12 genes.Bioinformatics analysis showed that the full length of SlJMJ2 was 2766 bp,encoding921 amino acids;Analyzing the secondary structure of a protein reveals that it containsα-Spiral and β-Corners,extended chains,and irregular curls,with more irregular curls and unstable hydrophobic proteins.SlJMJ12 has a total length of 1041 bp and encodes346 amino acids.Its protein secondary structure is characterized by more irregular curls and is a stable hydrophobic protein.(2)Fluorescence quantitative PCR results showed that SlJMJ2 and SlJMJ12 genes were expressed to varying degrees in different tissues of tomato.Among them,SlJMJ2 had the highest expression level in the old roots of tomato and the lowest expression level in the main roots.However,SlJMJ12,unlike SlJMJ2,had the highest expression level in young leaves and the lowest expression level in flowers.(3)The expression levels of SlJMJ2 and SlJMJ12 in tomato roots and leaves under Cd(cadmium),Hg(mercury),Cu(copper),and Pb(lead)stress were analyzed.The expression levels of SlJMJ2 in tomato roots and leaves showed a downward trend under Hg and Cu stress,an upward trend under Cd and Pb stress,and an upward trend under Hg and Pb stress.The expression of SlJMJ12 in roots and leaves was upregulated under Cd,Hg,Cu,and Pb stress.(4)Successfully constructed plant overexpression vectors SlJMJ2 and SlJMJ12.After performing Cd stress on transgenic Arabidopsis and wild type Arabidopsis,it was found that the germination rate and root length of overexpressed Arabidopsis were higher than those of wild type,while the root length and germination of mutant plants were lower than those of wild type,with significant differences.The mutants of SlJMJ2 and SlJMJ12 transgenic Arabidopsis and wild type Arabidopsis were stained with N-blue tetrazole(NBT).The mutant Arabidopsis was stained the deepest and the overexpressed Arabidopsis was stained the shallowest.After cadmium stress,the activities of SOD,POD,CAT in overexpressed Arabidopsis increased,the content of MDA decreased,and the content of chlorophyll was higher.(5)This article explores metal transporter related genes(IRT1,ZIP1,ABCC,NARPM4)and hormone related genes,including SL(D14,BRC1,CCD7),EBR(BIN2,BES1,BAK1),and ABA(ABI2,ABI5,ABF2).The results showed that metal transporter related genes and hormone related genes were elevated and higher in SlJMJ2 and SlJMJ12 overexpressions in Arabidopsis than in wild type,and the expression level of related genes in mutants was lower than that in wild type.