Studies On ZmCOP? And SAD Related To Oil Content In Maize Kernel

High-oil maize that results from a long-term artificial selection has important values in cooking oil,animal feed and bio-energy.Moreover,high-oil maize is a unique resource for genomics and genetics research.So identifying the genes associated with maize kernel oil content and validating their function will provide the gene resource and theoretical guidance for molecular breeding of high oil maize.In this study,association analysis,expression analysis,transgenic study were applied in investigating the function of coat protein complex ? gene?ZmCOP??and stearoyl-ACP desaturase genes?SAD?in maize.The major results are as follows:1.ZmCOPII encodes a coat protein complex ? which mediates the transport of proteins and lipids from the endoplasmic reticulum?ER?to the Golgi complex in early secretion pathway.In our GWAS results,a SNP in the second exon of ZmCOP? significantly associated with kernel oil concentration?P-3.4×10-14,n=368?.We then resequenced the region of ZmCOP? in a panel of 155 maize inbred lines and the association analysis based on ZmCOPII indicated that two insertion and/or deletion and 67 SNPs were significantly associated with maize kernel oil content.The linkage disequilibrium?LD?analysis indicated that two polymorphic sites in complete LD,InDel₂0 in 5'UTR and SNP₂988?Arginine/Cysteine?in the third exon,might be the functional sites for ZmCOPII,which were further validated in a panel of 508 maize inbred lines.A By855/Zheng58 F2 population and a B73/By804 RIL population that segregated at InDel₂0 and SNP₂988 were used for linkage analysis.The results indicated that these two sites were significantly associated with oil content.Whereas,the InDel₂0 did not associate with the expression of ZmCOPII,indicating that the SNP₂988 seems to be the functional site of ZmCOP?.In order to further validate the function of ZmCOPII,we transformed ZmCOP? into Arabidopsis mutants with T-DNA insertion in homologous gene,and found that the decreased seed oil content had been rescued indicating that ZmCOP? gene were associated with kernel oil content.2.Stearoyl-ACP desaturase?SAD?is a key rate-limiting enzyme for the conservation of stearic?C18:0?to oleic?C18:1?acid.In this study,11 SAD genes were identified in maize.Phylogenetic analysis releveled that SAD genes in maize have more divergent functions than Arabidopsis SAD genes.The genomic regional associations in a maize panel including 508 inbred lines identified 6 SAD genes significantly associated with the C18:0/C18:1 ratio or the level of C18:0 or C18:1?P<0.01?,one gene of which co-localized with a quantitative trait locus?QTL?and 5 of which co-localized with an expression QTL.Among these six SAD genes,ZmSADl located on chromosome 3 had the largest effect on C18:0/C18:1?P=1.17×10-4?,and localized with a QTL?LOD=11.2?and an expression QTL?P=1.58×10-14?.One nonsynonymous single-nucleotide polymorphism in exon 3 and one 5-bp insertion/deletion in the 3'UTR were further shown to contribute to the natural variation in C18:0/C18:1 according to ZmSAD1-based association mapping.These results will provide the theoretical guidance for the genetic improvement in maize kernel oil concentration and quality.