Imprinting,Methylation And Expression Characterization Of Maize EIN2-like

Gene imprinting is a way of the kinship effect that leads to different phenotypes of the parental by relying on single allele expression allele.Except for a few exceptions,most imprinted genes occur in the endosperm.As the most importants crops Maize in China,the level of Maize yield has a significant impact on Agriculture and food security.Grain is the main component of corn production.As a biological and economical storage organ,Maize endosperm plays an essential role in support of embryogenesis and seed germination,and provides abundant resource for human food and industrial products.Thus,identifying the imprinting genes and elucidating their functions in regulating endosperm development is of great significance for maize genetics and breeding.In the study,we cloned a maternally expressed gene ETHYLENE-INSENSITIVE 2-like(EIN2-like,GRMZM2G370991).We also analyzed its expression,the imprinting pattern in embryo and endosperm,and the methylation of endosperm DNA.in grain.In addition,we also explored its function of grain development in overexpressed EIN2-like gene of A.thaliana.The main results of this study are as follows:(1)The coding region of EIN2-like gene was 2019bp(without stop codon),encoding a total of 673 amino acids.The predicted protein was similar to that of EIN2 from Oryza sativa and Arabidopsis.(2)The qRT-PCR results showed that EIN2-like was highly expressed in maize kernel,especially in the endosperm,whereas low expressed in maize vegetative and reproductive organs(stem,root,leaf,ear,tassel).(3)We analyzed EIN2-like expression characteristic at later endosperm development stage(10-28 DAP)from B73×Mo17 and Mo17×B73.The qRT-PCR results showed that EIN2-like was specifically expressed in maize endosperm and was highly expressed at cell differentiation stage(8-12 DAP)in hybrid endosperm.(4)EIN2-like was maternally expressed both in endosperm and embryo.And the pattern of maternal expression remained unchanged at seed developmental stages(10-28DAP).(5)DNA methylation analysis showed that EIN2-like gene showed high methylation at the symmetric methylation site(CG and CHG)at the upstream region,while the maternal transcriptional transcript showed low methylation at the CG site in the upstream region and highly methylation at the CHG site.(6)We concluded that parental alleles of EIN2-like were differentially methylated in the symmetrical contexts and a DMR was identified at the 5’ upstream region of EIN2-like.(7)Overexpressed EIN2-like genes increased thousand-seed weight in Arabidopsis.But other traits included germination rate,biomass weight,photosynthetic pigment content were not significant between the transgenic lines and the wild type.These results indicated that maternally expressed gene EIN2-like might play crucial roles in regulation of seed development.