Identification And Isolation Of The Candidate Genes For Rscmv2 To Sugarcane Mosaic Virus In Maize

Corn is one of the most important food crops and feed resources.It can be affected by a variety of pathogens during the whole growth period and can't grow normally development,which affects the yield and quality of corn seriously.Maize(Zea mays L.)dwarf mosaic disease which is caused by sugarcane mosaic virus(SCMV)is a damaging viral disease occuring common in the world.It is the most basic way that is breeding and planting maize dwarf Mosaic disease resistant varieties to control this disease.On the basis of screening high quality resistant resources,mining and utilization of resistance genes is a precondition of cultivating disease-resistant varieties.In our previous study,using the line Siyi(completely resistant to SCMV)and Mo17(highly susceptible to SCMV)as donor parent and recurrent parent,respectively,near-isogenic lines(NILs)segregating at Rscmv2,but not the Rscmv1 region were developed to fine-map Rscmv2 to an interval of 196.5 kb.And two candidate genes named Zm ABP1 and Zm GAP for Rscmv2 were predicted,which encoding an auxin-binding protein 1 and a Rho GTPase-activating protein,respectively.On the basis of the above result,the main research content as follows:1.The 504 natural population which had been sequenced were planted in the Scientific &Educational Park of Henan Agricultural University in summer,2013.The 504 corn materials were conducted by inoculation,investigation,and using 556,000 SNP molecular markers for genome-wide association mapping of the major loci Rscmv2.We found five significant SNPs associated with resistance to SCMV in mean rating(P?1.796×10-6).Further more,we found five candidate genes though the publicly available B73 annotated genome.One of the five candidate genes GRMZM2G116204 can be detected in rating 1~3 stably,and the auxin binding protein 1encoded by GRMZM2G116204 related to plant defense,and the gene was located within the196.5Kb of fine-mapped Scmv2 region in our previous linkage analysis.So,the candidate gene GRMZM2G116204 encoding auxin binding protein 1 namely Zm ABP1 most likely is the target resistant gene we are interested in.2.We used the line Siyi(completely resistant to SCMV),Mo17(highly susceptible to SCMV)and the NILs Mo17AAbb(dan 3),Mo17 aa BB(dan 6),Mo17AABB(3+6)as materials in 2013,and used siyi,Mo17 as materials in 2014.The corn materials were conducted by inoculation,investigation,sampling,and application of real-time PCR technology to detect the relative expression of the two candidate genes Zm ABP1 and Zm GAP in resistant and susceptible lines.We found that the candidategene Zm ABP1 on the response of the virus SCMV is much higher than the candidate gene Zm GAP.Moreover,the results of phenotype identification show effectiveness of identification and accuracy of experimental materials.3.In order to further identify the function of the candidate gene Zm ABP1,we used the line siyi and Mo17 as materials and made the following experiments.we analyzed the coding sequences,introns and promoter sequences of the gene Zm ABP1 in the line Siyi and Mo17 by homology-based cloning.We find that the amino acid sequences are consistent between Siyi and Mo17,but the introns and promoter sequences have large variations which may be an important factor causing the disease.Furter more,the evolution analysis of amino acid sequences were carried out basing on protein sequences of the gene Zm ABP1,which provided a favorable basis for further understanding its function.In addition,a expression vector of the gene Zm ABP1 was constructed basing on it's ORF sequences,which can be used for functional verification by GM technology.