Dataset Information

De novo leaf and root transcriptome analysis identified novel genes involved in steroidal sapogenin biosynthesis in Asparagus racemosus.

ABSTRACT:

Background

Saponins are mainly amphipathic glycosides that posses many biological activities and confer potential health benefits to humans. Inspite of its medicinal attributes most of the triterpenes and enzymes involved in the saponin biosynthesis remains uncharacterized at the molecular level. Since the major steroidal components are present in the roots of A. racemosus our study is focussed on the comparative denovo transcriptome analysis of root versus leaf tissue and identifying some root specific transcripts involved in saponin biosynthesis using high-throughput next generation transcriptome sequencing.

Results

After sequencing, de novo assembly and quantitative assessment, 126861 unigenes were finally generated with an average length of 1200 bp. Then functional annotation and GO enrichment analysis was performed by aligning all-unigenes with public protein databases including NR, SwissProt, and KEGG. Differentially expressed genes in root were initially identified using the RPKM method using digital subtraction between root and leaf. Twenty seven putative secondary metabolite related transcripts were experimentally validated for their expression in root or leaf tissue using q-RT PCR analysis. Most of the above selected transcripts showed preferential expression in root as compared to leaf supporting the digitally subtracted result obtained. The methyl jasmonate application induces the secondary metabolite related gene transcripts leading to their increased accumulation in plants. Therefore, the identified transcripts related to saponin biosynthesis were further analyzed for their induced expression after 3, 5 and 12 hours of exogenous application of Methyl Jasmonate in tissue specific manner.

Conclusions

In this study, we have identified a large set of cDNA unigenes from A. racemosus leaf and root tissue. This is the first transcriptome sequencing of this non-model species using Illumina, a next generation sequencing technology. The present study has also identified number of root specific transcripts showing homology with saponin biosynthetic pathway. An integrated pathway of identified saponin biosynthesis transcripts their tissue specific expression and induced accumulation after methyl jasmonate treatment was discussed.

SUBMITTER: Upadhyay S

PROVIDER: S-EPMC4162912 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Asparagus racemosus is known for its diverse content of secondary metabolites, i.e., saponins, alkaloids, and a wide range of flavonoids. Flavonoids, including phenols and polyphenols, have a significant role in plant physiology and are synthesized in several tissues. Despite the diverse role of flavonoids, genetic information is limited for flavonoid biosynthesis pathways in A. racemosus. The current study explores full-scale functional genomics information of A. racemosus by de novo transcriptome sequencing using Illumina paired-end sequencing technology to elucidate the genes involved in flavonoid biosynthesis pathways. The de novo assembly of high-quality paired-end reads resulted in ∼2.3 million high-quality reads with a pooled transcript of 45,647 comprising ∼76 Mb transcriptome with a mean length (bp) of 1,674 and N50 of 1,868bp. Furthermore, the coding sequence (CDS) prediction analysis from 45,647 pooled transcripts resulted in 45,444 CDS with a total length and mean length of 76,398,686 and 1,674, respectively. The Gene Ontology (GO) analysis resulted in a high number of CDSs assigned to 25,342 GO terms, which grouped the predicted CDS into three main domains, i.e., Biological Process (19,550), Molecular Function (19,873), and Cellular Component (14,577). The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway database was used to categorize 6,353 CDS into 25 distinct biological pathway categories, in which the majority of mapped CDS were shown to be related to translation (645), followed by signal transduction (532), carbohydrate metabolism (524), folding, sorting, and degradation (522). Among these, only ∼64 and 14 CDSs were found to be involved in the phenylpropanoid and flavonoid biosynthesis pathways, respectively. Quantitative Real-time PCR was used to check the expression profile of fourteen potential flavonoid biosynthesis pathway genes. The qRT-PCR analysis result matches the transcriptome sequence data validating the Illumina sequence results. Moreover, a large number of genes associated with the flavonoids biosynthesis pathway were found to be upregulated under the induction of methyl jasmonate. The present-day study on transcriptome sequence data of A. racemosus can be utilized for characterizing genes involved in flavonoid biosynthesis pathways and for functional genomics analysis in A. racemosus using the reverse genetics approach (CRISPR/Cas9 technology).