Molecular cloning, sequence analysis, and construction of expression vector of monofunction feedback inhibition sensitive CtAK1 gene in Carthamus tinctorius / 中草药

ABSTRACT

Objective: To separate the full length cDNA of gene encoding the key enzyme AK in aspartate metabolic pathways of Carthamus tinctorius and to construct plant overexpression vector. Methods: According to annotation on transcriptome library of C. tinctorius and core fragments and expression analysis data of CtAK identified by qRT-PCR, we separated the full length cDNA of AK gene of C. tinctorius (CtAK) using RACE technology and constructed plant expression vector using recombinant DNA technology. Results: Bioinformatics analysis showed the full length CtAK was 1 703 bp and ORF area was 1 626 bp, encoding 541 amino acid residues. The function structure domain analysis showed the gene might be a monofunction feedback inhibition sensitive AK1 from plant. We successfully constructed plant expression vector pCAMBIA3301-CTP-AK1 which contained 35 S promoter and Bar resistance genes and chloroplast transit peptide by recombinant DNA technology. Conclusion: The gene encoding CtAK1 is obtained and the plant overexpression vector is constructed, which lays the foundation for researching on biological function and mechanism of action in amino acid metabolism regulation of C. tinctorius.