Sequence and expression analyses of three seed protein genes of buckwheat

Objective To clarify the sequence features and functions of three seed protein genes of buckwheat, providing a theoretical foundation for studying the molecular mechanism during seed development.

Method Using local Perl script, BLASTP, MEGA6 and PLANTCARE, the sequences and evolutionary relationships of three buckwheat seed protein genes, namely 13S globulin gene, 11S globulin gene and glutelin gene, were analyzed. The cis-regulatory elements in the promoter region were predicted. Using the qRT-PCR technology, the expression patterns of three genes were analyzed during seed development of Fagopyrum esculentum, F. tataricum and F. cymosum.

Result The 13S globulin gene included four exons and three introns. The protein length was 515 aa. This 13S globulin only had six different amino acid sites compared with another 13S globulin in F. tataricum (ABI32184.1), thus they might be the same gene. The expression level was the highest in the middle filling stage of F. tataricum. The 11S globulin gene included eight exons and seven introns. The protein length was 914 aa, and it was most closly related to a 11S globulin in Sagittaria sagittifolia (CAA70334.1). The expression level was the highest in the middle filling stage of F. tataricum. The glutelin gene included three exons and two introns. The protein length was 355 aa, and it was most closly related to a B5-type glutelin in Beta vulgaris (XP_010683769.1). The expression level was the highest in the middle filling stage of F. esculentum. The promoter regions of three genes contained various cis-acting elements which responded to stresses and phytohormones, as well as cis-acting elements required for endosperm expression.

Conclusion The sequences of 13S globulin, 11S globulin and glutelin genes are obtained. These genes might be preferentially expressed in endosperm, and involved in stress and hormone responsive processes. The middle filling stage is the key stage for seed protein accumulation, and three genes are massively expressed at this stage, which provides a solid foundation for studying gene function in the future.