Cloning，Functional Sequence Analysis and Prokaryotic Expression of Cotton GhAO3 cDNA

Abstract

Abstract: A cotton ascorbate oxidase(GhAO3)full-length open reading frame（ORF）cDNA was cloned from elongating fiber tissue（15 DPA）by RT-PCR method, GhAO3 ORF contains 1 758 bp nucleotides and codes a protein of 585 amino acids. Through sequence function analysis and homology sequence alignment, GhAO3 protein includes three multi-copper oxidase domain and transmembrane signal sequence with a high conservation. Phylogenetic tree analysis showed that GhAO3 has a similar relationship with GmAO protein. Prokaryotic expression vector pET32a-GhAO3 was constructed and transformated into E. coli BL21（DE3）. Recombinant GhAO3 protein was obtained after induction by IPTG and SDS-PAGE analysis with a MW of 64 kD, and had a higher enzymatic activity.

Key words: Cotton ascorbate oxidase, Gene cloning, Functional sequence analysis, Prokaryotic expression

Ran Maofei, He Yi, Qian Wenjie, Wang Fei, Li Hongbin. Cloning，Functional Sequence Analysis and Prokaryotic Expression of Cotton GhAO3 cDNA[J]. Biotechnology Bulletin, 2013, 0(8): 51-56.

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