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Cloning and Characterization of an Agglutinin Gene from Arisaema lobatum
Authors:Juan Lin  Xuanwei Zhou  Yongzhen Pang  Han Gao  Jiong Fei  Guo-an Shen  Jin Wang  Xinsheng Li  Xiaofen Sun  Kexuan Tang
Institution:(1) State Key Laboratory of Genetic Engineering, School of Life Sciences, Morgan-Tan International Center for Life Sciences, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center, Fudan University, Shanghai, 200433, P.R. China;(2) Plant Biotechnology Research Center, Fudan-SJTU-Nottingham, Plant Biotechnology R&D Center, School of Agriculture and Biology, Shanghai Jiao Tong University, Shanghai, 200030, P.R. China;(3) Shaanxi Key Laboratory of Bio-resources, Shaanxi University of Technology, Hanzhong, 723000, P.R. China
Abstract:A novel agglutinin gene was cloned from Arisaema lobatum using SMART RACE-PCR technology. The full-length cDNA of Arisaema lobatum agglutinin (ala) was 1078 bp and contained a 774 bp open reading frame encoding a lectin precursor (proproprotein) of 258 amino acid residues with a 23 aa signal peptide. ALA contained three mannose-binding sites (QXDXNXVXY) with two-conserved domains of 45% identity, ALA-DOM1 and ALA-DOM2. The three-dimensional structure of ALA was very similar to that of GNA (Galanthus nivalis agglutinin). ALA shared varying identities, ranging from 40% to 85%, with mannose-binding lectins from other species of plant families, such as Araceae, Alliaceae, Iridaceae, Lillaceae, Amaryllidaceae and Bromeliaceae. Genomic sequence of ala was also cloned using genomic walker technology, and it was found to contain three putative TATA boxes and eight possible CAAT boxes in the 5′-flanking region. No intron was found within the region of genomic sequence. Southern blot analysis indicated that the ala belonged to a multi-copy gene family. Expression pattern analysis revealed that the ala preferentially expressed in the tissues with the higher expression being found in spadix, bud, leaf, spathe and tuber. The cloning of the ala gene not only provides a basis for further investigation of its structure, expression and regulation mechanism, but also enables us to test its potential role in controlling pests and fungal diseases by transferring the gene into plants in the future.
Keywords:Arisaema lobatum  Arisaema lobatum agglutinin (ALA)  cDNA cloning  genomic walker technology  mannose-binding lectin  RACE
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