Cloning and characterization of an agglutinin gene from Arisaema lobatum.
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.
Lin
Juan
J
State Key Laboratory of Genetic Engineering, School of Life Sciences, Morgan-Tan International Center for Life Sciences, Fudan University, Shanghai, 200433, PR China.
Zhou
Xuanwei
X
Pang
Yongzhen
Y
Gao
Han
H
Fei
Jiong
J
Shen
Guo-an
GA
Wang
Jin
J
Li
Xinsheng
X
Sun
Xiaofen
X
Tang
Kexuan
K
D000373:Agglutinins / Q000737:chemistry / Q000235:genetics*; D000595:Amino Acid Sequence; D031057:Arisaema / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites; D015139:Blotting, Southern; D003001:Cloning, Molecular; D004247:DNA / Q000737:chemistry; D017931:DNA Primers / Q000737:chemistry; D018076:DNA, Complementary / Q000378:metabolism; D019143:Evolution, Molecular; D018506:Gene Expression Regulation, Plant; D015723:Gene Library; D017343:Genes, Plant; D018745:Genome, Plant; D037102:Lectins / Q000737:chemistry; D008358:Mannose / Q000737:chemistry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010802:Phylogeny; D016133:Polymerase Chain Reaction; D021382:Protein Sorting Signals; D017434:Protein Structure, Tertiary; D012313:RNA / Q000737:chemistry; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid
D000373:Agglutinins; D017931:DNA Primers; D018076:DNA, Complementary; D037102:Lectins; D021382:Protein Sorting Signals; D012313:RNA; D004247:DNA; D008358:Mannose