Cloning and expression of a novel cDNA encoding a mannose-binding lectin from Dendrobium officinale.
Using RNA extracted from Dendrobium officinale young leaves and primers designed according to the conservative regions of orchidaceae lectins, the full-length cDNA of Dendrobium officinale agglutinin2 (DOA2) was cloned by rapid amplification of cDNA ends (RACE). The full-length cDNA of doa2 was 777 bp and contained a 513 bp open reading frame (ORF) encoding a lectin precursor of 170 amino acids. Through comparative analysis of doa2 gene and its deduced amino acid sequence with those of other orchidaceae species and Amaryllidaceae species, it was found that DOA2 had many common characters of mannose-binding lectin superfamily including three mannose-binding sites. Semi-Quantitative RT-PCR analysis revealed that doa2 mRNA expression was detected in all tested tissues including root, stem and leaf, however, the expression was higher in stem, and lower in leaf. As the doa2 mRNA was detected in all the tested plant tissues, the doa2 was considered to be a constitutively expressed gene. The recombinant protein was expressed in E. coli and purified. Anti-fungal assay showed that DOA2 has anti-fungal activity towards Gibberella zeae. To our knowledge, this is the first report on cDNA cloning of mannose binding lectin from Dendrobium officinale.
Chen
Zhonghai
Z
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, People's Republic of China. zh10246@163.com
Sun
Xiaofen
X
Tang
Kexuan
K
D000373:Agglutinins / Q000235:genetics*; D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D002681:China; D003001:Cloning, Molecular; D017931:DNA Primers; D018076:DNA, Complementary / Q000235:genetics; D031667:Dendrobium / Q000235:genetics*; D037601:Mannose-Binding Lectin / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D021141:Nucleic Acid Amplification Techniques; D012333:RNA, Messenger / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013045:Species Specificity
D000373:Agglutinins; D017931:DNA Primers; D018076:DNA, Complementary; D037601:Mannose-Binding Lectin; D012333:RNA, Messenger