Molecular cloning of a lectin cDNA from Alocasia macrorrhiza and prediction of its characteristics.
The cDNA of Alocasia macrorrhiza lectin (aml, GenBank accession number: DQ340864) was cloned by RACE-PCR and its characteristics were predicted by various bioinformatics tools. GSPs (Gene Specific Primers) were designed according to the conserved regions of the genes encoded for lectins and similar proteins from the same family Araceae. Total RNAs were extracted from the tubers of A macrorrhiza by Qiagen RNeasy mini kit. The 3'- and 5'-RACE-PCRs were performed with the isolated total RNAs by SMART(TM)RACE cDNA amplification kit from BD Biosciences Clontech Company, respectively. The purified PCR products were ligated with pMD 18-T vector, and the confirmed clones were sequenced. The full-length cDNA of aml was obtained by combination of 3'- and 5'-end sequences, and was then confirmed by full-length 3'-RACE-PCR. The aml cDNA is 1 124 bp long. The deduced amino acid length of AML lectin is 270 aa. Its relative molecular weight is 29.7 kD. The results of homologous analysis showed a high similarity between AML and other mannose-binding lectins and similar proteins from Araceae family. Two typical B-lectin domains and three mannose- binding motifs were found in the sequence of AML. With all these taken together, it can be concluded that this newly cloned aml cDNA encodes for a mannose-binding lectin.
Zhu
Ya-Ran
YR
College of Natural Resources and Environment, College of Life Sciences, South China Agricultural University, Guangzhou 510642, China.
Wang
Jie
J
Huang
Bing-Qiu
BQ
Hou
Xue-Wen
XW
D031055:Alocasia / Q000235:genetics*; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D019295:Computational Biology; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010802:Phylogeny; D037121:Plant Lectins / Q000737:chemistry / Q000145:classification / Q000235:genetics*; D017433:Protein Structure, Secondary; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D037121:Plant Lectins