Molecular cloning, characterization, and expression analysis of lignin biosynthesis genes from kenaf (Hibiscus cannabinus L.)

Abstract

Kenaf (Hibiscus cannabinus L.) is an annual herbaceous crop and the third largest fiber crop of economic importance, after cotton and jute. In view of recent global environmental challenges, multipurpose uses of kenaf have been developed, such as animal feed, bio-plastics, bio-fuels, and phytoremediation. We cloned genes related to monolignol biosynthesis using a reverse transcriptase PCR (RT-PCR) strategy. ‘Auxu’ cultivars in the seedling phase were used as the experimental source material. The 4-coumarate:CoAligase (4CL), cinnamoyl-CoA reductase (CCR), ferulate 5-hydroxylase (F5H), caffeic acid O-methyltransferase (COMT), cinnamyl alcohol dehydrogenase (CAD), encoding hydroxycinnamoyl transferase (HCT), ρ-coumarate 3-hydroxylase (C3H) and caffeoyl-CoA-O-methyltransferase (CCoAOMT), cDNAs had open reading frames of 1623, 1017, 1179, 921, 1107, 1323, 1527 and 744 bp, encoding polypeptides of 540, 338, 392, 306, 368, 441, 508 and 247 amino acid residues, respectively. We submitted the five gene sequences (4CL, CCR, F5H, COMT, and CAD) to the GenBank database. A phylogenetic tree showed that lignin biosynthesis genes had the closest relationships with Gossypium and Theobroma cacao. Expression levels of the genes 4CL, HCT, C3H, CCoAOMT, CCR, F5H, COMT, and CAD in stems from plants harvested at four different growth stages were evaluated. Expression levels of 4CL, HCT, C3H, CCoAOMT, CCR, F5H, COMT, and CAD were gradually increased during the vegetative growth of kenaf stem. Similarly with the gene expression, lignin amounts were also increased during the stem growth. Thus, these six genes may be directly involved in lignin biosynthesis. These results will be useful basic data for molecular breeding to enhance bio-mass production.