Genome-Wide Identification, Classification, Expression Profiling, and SSR Marker Development of the MADS-Box Gene Family in Citrus

MADS-box genes encode transcription factors that are involved in developmental control and signal transduction in plants, animals and fungi. To date, few citrus MADS-box genes and limited tissue expression profiling have been reported. In this study, 84 clementine mandarin and 52 sweet orange MADS-box genes have been identified using bioinformatics analysis. Subsequently, comparative analysis of these genes, including gene structures, phylogenetic relationships, conserved protein motifs, expression patterns, and simple sequence repeat (SSR) markers were identified in these citrus species. According to these analyses, these 136 MADS-box genes can be categorized into five (MIKCC, MIKC*, M alpha, M beta, and M gamma) groups, and the MIKCC group was further divided into 13 subfamilies in two citrus species. In addition, gene duplication of MADS-box genes was also investigated in the clementine mandarin, sweet orange, rice and Arabidopsis genomes. A lower percentage of MADS-box gene duplication was found in the clementine mandarin and sweet orange genomes as compared with Arabidopsis. A survey of the two citrus species based on organ-specific expression profiling indicated that MADS-box genes exhibit various expression patterns, suggesting diverse and novel functions of MADS-box gene families in citrus. In addition, 128 new putative microsatellites with flanking sequences sufficient for primer design were also identified from 136 MADS genes. Sixteen SSRs were selected randomly for validation and transferability testing in 11 citrus species. Among 16 SSRs, 15 were identified as true-to-type SSR loci and were highly transferable among citrus species. These results provide a useful reference for the selection of candidate MADS-box genes for cloning and further functional analysis in citrus.