Molecular Cloning And Functional Analysis Of GMCINV1and GMSPP1Genes Involved In Sucrose Accumulation From Soybean

Vegetable soybean is a special soybean [Glycine max (L.) Merr.], harvested in the R6-R7growth stage with full seeds and still green pods. Sucrose content is important factor of eating quality for vegetable soybean. In this study, GmSPPl and GmCInv1were cloned and analyzed the function, both involved in sugar metabolism. It was useful for better understand the impact of the two genes on sugar accumulation and plant growth and development. The results are following:1.Detection of the sucrose content in different tissues of soybeanThe sucrose content in the fresh soybean seeds keep increasing in the early seed development stage. And the sucrose content in the root and flower is higher in the flower development stage.2.Gene isolation and sequence analysisWe have isolated GmCInvl gene from soybean by gene homology, using CInv1sequence of Arabidopsis. GmCInv1contained a1668bp open reading frame, encoding a555amino acids protein (63.20kD). The phylogenetic analysis indicated that GmCInv1protein has more relationship to Ljinvl, and both showed92%sequence identity.3. Expression pattern analysis and functional analysis of GmCInv1transgenic Arabidopsis thalianaReal-time Quantitative PCR was used to analyze the expression pattern of GmCInvl in soybean. GmCInv1gene was all detected in different tissues, but transcription level was different, GmCInv1strongly expressed in roots and flowers. During leaf development, GmCInv1weakly expressed in young leaves, but strongly expressed in old leaves. Transformants with overexpression GmCInvl displayed a range of phenotypes. The transgenic plants in the leaves had low levels of sucrose but contained high glucose than control, However, there are no significant difference in seeds and delay in flowering. The proline content of GmCInv1in transgenic Arabidopsis thaliana was higher than wild-type. 4. Cloning and sequence analysis of GmSPP1geneWe have isolated GmSPP1gene from soybean by gene homology, using AtSPP1sequence of Arabidopsis. GmSPP1contains a1257bp open reading frame, encoding a418amino acids protein (47.22kD). The phylogenetic analysis showed that GmSPP1protein has more relationship to that of grapes.5. Expression pattern analysis and functional analysis of GmSPP1GmSPP1expression levels increase throughout seed development. We found that the expression levels of GmSPP1fluctuated in LD and LL with approximately24h in LD and12h in LL. We observed a correlation between sucrose content and GmSPP1expression levels. GmSPP1transgenic lines have not significant differences in sucrose content compared with WT in the leaves, but there are significant difference in the seeds.