Functional genomics studies of costunolide biosynthesis in Saussurea lappa

Abstract

Saussurea lappa has innumerable pharmacological properties and sesquiterpene lactones are considered as the principle components confering these properties to the plant. Costunolide is among the chief sesquiterpene lactones occurring in the plant. De novo transcriptome sequencing of leaf tissue using Illumina HiSeq 2000 sequencing platform yielded 62,039,614 high quality paired end raw reads. These reads were assembled using Trinity assembler and 122,434 transcripts were obtained which were further clustered into 110,283 unigenes using CD-HIT. The transcripts were functionally annotated using BLAST similarity search tool against several databases. The transcripts showed significant homology with already known genes. Further, about newline9,536 GO terms were assigned to the transcripts, which were classified into three categories viz. molecular function, biological process and cellular component category. The SSR markers were discovered in the transcripts using MISA perl script and 9.05% transcripts were found to contain SSRs. In-silico analysis enabled the discovery of mevalonate pathway and costunolide biosynthetic genes. Also, genes involved in flavonoid and alkaloid biosynthesis were obtained in the dataset. Further qRT-PCR based relative expression analysis was conducted and higher expression of genes of costunolide biosynthesis was observed in leaf tissue as compared to root tissue. The molecular cloning of Mevalonate kinase, Phosphomevalonate kinase, Diphosphomevalonate decarboxylase and Germacrene A Synthase was also carried out. This is the pioneer study pertaining to the functional genomics studies in S. lappa.