Cloning And Functional Characterization Of The Sex Determination Gene TASSELSEED5 In Maize

Maize is one of the most important food and energy crop in the world.Its offspring has a closed relationship with floret organs.At the same time,maize is bisexual grass plants.The mature male and female florets form in tassel and ear respectively.Thus as a model plant,maize is used to study flower development.Many tasselseed mutants have a reversal in sex determination,which leads to the generation of kernals in tassel eventually,such as two recessive mutants,tasselseed1(ts1)and tasselseed2(ts2).Intriguingly,we mapped the dominant Tasselseed5(Ts5)mutant whose phenotype is similar with ts2.The phenotype of Ts5 mainly appears in tassel and ear,especially in the progress of floret formation.With the development of tassel spikelet,pistil is arrested in wide type,while the floret in tassel can produce pistil in Ts5.Each ear spikelet consists of an upper and lower floret enclosed by a pair of gulmes.The lower floret and the stamens in the upper floret degenerate.Only the upper floret matures produce kernals,while many lower florets can produce kernals in Ts5.We mapped the TASSELSEED5 gene and confirmed that TS5 was located in the 0.6 Mb internal,positioning in the short arm of chromosome 4.There are 7 protein coding genes in this region.We checked the expression level of 7 genes in the tassel when the florets were in sex determination period.The results showed that only GRMZM2G177668 gene had a high expression level in mutants while others not.Blasting the protein sequence,we found a homologous gene in Arabidopsis thaliana,and its number is AT3G48520.This gene codes a cytochrome C oxidase,which can transform JA-Ile to 120H-JA-Ile.Thus,we test the hormone level of tassel in wide type and mutant.The results indicated that there was no peak in mutant tassel during sex determinate period compared with wide type.Meanwhile,we applied the JA to developing mutant tassel,and found that JA could rescue the wide type phenotype to a certain level.In situ hybridization assay showed us the pattern of gene expression.It mainly expressed in pistil stem cells,and the expression pattern of TS5 was similar with TS2.Besides that,qRT-PCR analysis showed that TS5 was expressed in root,stem,leaf,tassel and ear.More important,the gene expression level of TS5 in mutant was always higher than that in wide type.In conclusion,the phenotype of Ts5 mutant was due to the higher expression level of GRMZM2G177668 in special space and times.Its cloning and functioning analysis widen our understanding in sex determination.Meanwhile,the extra kernals in ear of Ts5 mutant is expected to increase production.