Cloning and prokaryotic expression analysis of LcMADS12 from Leymus chinensis

doi:10.11686/cyxb2017323

Abstract

Abstract: In flowering plants, the E-function genes, which belong to the MADS-box gene family, play central roles in floral meristem and floral organ development. To understand the molecular mechanisms of floral development in Leymus chinensis, a MADS-box gene, LcMADS12, was cloned according to RNA-sequencing data obtained in this research. The LcMADS12 gene contained a 741 bp open reading frame (ORF) which encoded 246 amino acids. Bioinformatics analysis showed that the LcMADS12 encoded an amino acid sequence which did not include signal peptide and the transmembrane region. Multi-sequence alignment and functional domain analysis showed that LcMADS12 had a conserved MADS-box domain and a semi-conserved K region, which was a member of the E-class functional gene SEP subfamily. Phylogenetic analysis showed that LcMADS12 had a 99% homology with wheat TaAGL16, and 86% homology with OsMADS7. Tissue-specific analysis showed that no expression of LcMADS12 gene was detected in the vegetative organs (root, stem, leaf) of L. chinensis. In the floral organ, the expression of LcMADS12 was detected higher in the mature stamen, pistil and palea, and was lower or not expressed in lemma and glume. These results indicate that expression of the LcMADS12 gene is tissue-specific. We suggest that LcMADS12 could play an important role in the reproductive development of L. chinensis. A yeast one-hybrid experiment showed that LcMADS12 had a transcriptional activation function. In addition, a prokaryotic expression vector was constructed and the fusion protein could be expressed highly. The cloning of LcMADS12 and the acquisition of fusion proteins has laid the foundation for further study of the function of this gene.

Key words: Leymus chinensis, MADS-box transcription factor, LcMADS12, expressing analysis, Prokaryotic expression

JIA Jun-ting, ZHAO Pin-cang, LIU Zhu-jiang, YUAN Guang-xiao, YANG Wei-guang, LIU Shu, CHEN Shuang-yan, LI Xiao-xia, LIU Gong-she. Cloning and prokaryotic expression analysis of LcMADS12 from Leymus chinensis[J]. Acta Prataculturae Sinica, 2018, 27(7): 64-72.

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