Molecular Cloning of GhCAD6 Gene and Construction of Its Sense,Anti-sense and RNAi Expression Vectors

Cinnamyl alcohol dehydrogenase(CAD)(EC 1.1.1.195) catalyses the final step in lignin precursor synthesis reducing the cinnamyl aldehydes to the corresponding alcohols in the presence of NADPH.In this paper,we report the molecular cloning a length of 1 569 bp sequence from genomic DNA of GhCAD6 by PCR.The genomic DNA of GhCAD6 contains five exons and four introns.To investigate the function of GhCAD6 gene,the sense and anti-sense expression vectors of GhCAD6 were constructed,in which the coding region of the gene was placed under the E6 promoter in either sense or anti-sense orientation.In addition,especial 417 bp fragments of GhCAD6 NBD were also cloned,and this fragment was inserted into plant vector to construct RNAi expression vector of GhCAD6.These constructed vectors were then transformed into Agrobacterium LBA4404.These constructed vectors provided an effective tool for the further study of GhCAD6 gene function.