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Next Generation Sequencing and Transcriptome Analysis Predicts Biosynthetic Pathway of Sennosides from Senna (Cassia angustifolia Vahl.), a Non-Model Plant with Potent Laxative Properties.

Rama Reddy NR, Mehta RH, Soni PH, Makasana J, Gajbhiye NA, Ponnuchamy M, Kumar J - PLoS ONE (2015)

Bottom Line: A total of 10,763 CDS differentially expressing in both young and mature leaf libraries of which 2,343 (21.7%) CDS were up-regulated in young compared to mature leaf.Several differentially expressed genes found functionally associated with sennoside biosynthesis.We have identified a set of putative genes involved in various secondary metabolite pathways, especially those related to the synthesis of sennosides which will serve as an important platform for public information about gene expression, genomics, and functional genomics in senna.

View Article: PubMed Central - PubMed

Affiliation: ICAR-Directorate of Medicinal and Aromatic Plants Research (DMAPR), Anand, Gujarat, India.

ABSTRACT
Senna (Cassia angustifolia Vahl.) is a world's natural laxative medicinal plant. Laxative properties are due to sennosides (anthraquinone glycosides) natural products. However, little genetic information is available for this species, especially concerning the biosynthetic pathways of sennosides. We present here the transcriptome sequencing of young and mature leaf tissue of Cassia angustifolia using Illumina MiSeq platform that resulted in a total of 6.34 Gb of raw nucleotide sequence. The sequence assembly resulted in 42230 and 37174 transcripts with an average length of 1119 bp and 1467 bp for young and mature leaf, respectively. The transcripts were annotated using NCBI BLAST with 'green plant database (txid 33090)', Swiss Prot, Kyoto Encylcopedia of Genes & Genomes (KEGG), Cluster of Orthologous Gene (COG) and Gene Ontology (GO). Out of the total transcripts, 40138 (95.0%) and 36349 (97.7%) from young and mature leaf, respectively, were annotated by BLASTX against green plant database of NCBI. We used InterProscan to see protein similarity at domain level, a total of 34031 (young leaf) and 32077 (mature leaf) transcripts were annotated against the Pfam domains. All transcripts from young and mature leaf were assigned to 191 KEGG pathways. There were 166 and 159 CDS, respectively, from young and mature leaf involved in metabolism of terpenoids and polyketides. Many CDS encoding enzymes leading to biosynthesis of sennosides were identified. A total of 10,763 CDS differentially expressing in both young and mature leaf libraries of which 2,343 (21.7%) CDS were up-regulated in young compared to mature leaf. Several differentially expressed genes found functionally associated with sennoside biosynthesis. CDS encoding for many CYPs and TF families were identified having probable roles in metabolism of primary as well as secondary metabolites. We developed SSR markers for molecular breeding of senna. We have identified a set of putative genes involved in various secondary metabolite pathways, especially those related to the synthesis of sennosides which will serve as an important platform for public information about gene expression, genomics, and functional genomics in senna.

No MeSH data available.


GO Classification in A) young and B) mature leaf transcriptome of Cassia angustifolia.Cassia angustifolia CDS were searched against the non-redundent protein sequences available in the Uni-ProtKB/SwisProt database using BLASTX with E value threshold of 1e-06 in order to assign putative function. Out of 42,230 and 37,174 CDS in young and mature leaf respectively, 29,944 (70.9%) CDS in young and 28,099 (75.5%) CDS in mature leaf transcriptome showed significant hits to the Uni-ProtKB/SwisProt data set thereby showing overall gene conservation. In addition, many C. angustifolia transcripts showed homology to uncharacterised proteins annotated as unknown, hypothetical and expressed proteins.
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pone.0129422.g002: GO Classification in A) young and B) mature leaf transcriptome of Cassia angustifolia.Cassia angustifolia CDS were searched against the non-redundent protein sequences available in the Uni-ProtKB/SwisProt database using BLASTX with E value threshold of 1e-06 in order to assign putative function. Out of 42,230 and 37,174 CDS in young and mature leaf respectively, 29,944 (70.9%) CDS in young and 28,099 (75.5%) CDS in mature leaf transcriptome showed significant hits to the Uni-ProtKB/SwisProt data set thereby showing overall gene conservation. In addition, many C. angustifolia transcripts showed homology to uncharacterised proteins annotated as unknown, hypothetical and expressed proteins.

Mentions: Gene ontology (GO) assignments were used to classify the functions of the predicted CDS. The GO mapping also provides ontology of defined terms representing gene product properties which were grouped into three main domains: Biological process, Molecular function, and Cellular component. GO terms were assigned for 25,337 and 22,975 annotated CDS in young and matured leaves, respectively (Table 3 and S6 Fig). As one GO term can be assigned to multiple CDS and the single CDS can have multiple GO terms, a total number of 53,278 and 48,466 GO terms were enriched for the annotated CDS in young and mature leaves, respectively. In young leaf, we have obtained 19,811 terms in biological process, 20,578 terms in molecular functions and 12,889 terms in cellular component (Table 3; S6 Fig). For mature leaf, 18,156 terms were mapped into biological process, 18,823 terms were mapped into molecular functions, and 11,487 terms were mapped into cellular component (Table 3; S6 Fig). The WEGO plots were plotted based on GO hits and CDS were categorized into 45 functional groups from WEGO analysis (Fig 2). In the biological process category, metabolic process (GO:0008152) (young 15391, 60.7% and mature 14395, 62.7%) followed by cellular process (GO:0009987) (young 13691, 54.0% and mature 12381, 53.9%) were prominent in young and mature leaves, suggesting these CDS might be involved in some important metabolic activities in senna. In the molecular function category, “catalytic activity” (GO: 0003824) (young 12555, 49.6% and mature 12049, 52.4%) represented most abundant term, followed by “binding activity” (GO: 0005488) (young 12513, 49.4% and mature 11394, 49.6%). Extremely low percentage of genes were classified in terms of “protein tag” (GO: 0031386), “locomotion” (GO: 0040011), “metallochaperone” (GO: 0016530) and “viral reproduction” (GO: 0016032) in young as well as mature leaf transcriptomes. Under the cellular component category, highest number of CDS was associated with “cell” (GO: 0005623) (young 12790, 50.5% and mature 11377, 49.5%) and “cell part” (GO: 0044464) (young 12790, 50.5% and mature 11377, 49.5%) followed by organelle (GO: 0043226) (young 7382, 29.1% and mature 6580, 28.6%) in young and mature leaf. Both the libraries showed similar type of distribution pattern of CDS under different GO terms.


Next Generation Sequencing and Transcriptome Analysis Predicts Biosynthetic Pathway of Sennosides from Senna (Cassia angustifolia Vahl.), a Non-Model Plant with Potent Laxative Properties.

Rama Reddy NR, Mehta RH, Soni PH, Makasana J, Gajbhiye NA, Ponnuchamy M, Kumar J - PLoS ONE (2015)

GO Classification in A) young and B) mature leaf transcriptome of Cassia angustifolia.Cassia angustifolia CDS were searched against the non-redundent protein sequences available in the Uni-ProtKB/SwisProt database using BLASTX with E value threshold of 1e-06 in order to assign putative function. Out of 42,230 and 37,174 CDS in young and mature leaf respectively, 29,944 (70.9%) CDS in young and 28,099 (75.5%) CDS in mature leaf transcriptome showed significant hits to the Uni-ProtKB/SwisProt data set thereby showing overall gene conservation. In addition, many C. angustifolia transcripts showed homology to uncharacterised proteins annotated as unknown, hypothetical and expressed proteins.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4476680&req=5

pone.0129422.g002: GO Classification in A) young and B) mature leaf transcriptome of Cassia angustifolia.Cassia angustifolia CDS were searched against the non-redundent protein sequences available in the Uni-ProtKB/SwisProt database using BLASTX with E value threshold of 1e-06 in order to assign putative function. Out of 42,230 and 37,174 CDS in young and mature leaf respectively, 29,944 (70.9%) CDS in young and 28,099 (75.5%) CDS in mature leaf transcriptome showed significant hits to the Uni-ProtKB/SwisProt data set thereby showing overall gene conservation. In addition, many C. angustifolia transcripts showed homology to uncharacterised proteins annotated as unknown, hypothetical and expressed proteins.
Mentions: Gene ontology (GO) assignments were used to classify the functions of the predicted CDS. The GO mapping also provides ontology of defined terms representing gene product properties which were grouped into three main domains: Biological process, Molecular function, and Cellular component. GO terms were assigned for 25,337 and 22,975 annotated CDS in young and matured leaves, respectively (Table 3 and S6 Fig). As one GO term can be assigned to multiple CDS and the single CDS can have multiple GO terms, a total number of 53,278 and 48,466 GO terms were enriched for the annotated CDS in young and mature leaves, respectively. In young leaf, we have obtained 19,811 terms in biological process, 20,578 terms in molecular functions and 12,889 terms in cellular component (Table 3; S6 Fig). For mature leaf, 18,156 terms were mapped into biological process, 18,823 terms were mapped into molecular functions, and 11,487 terms were mapped into cellular component (Table 3; S6 Fig). The WEGO plots were plotted based on GO hits and CDS were categorized into 45 functional groups from WEGO analysis (Fig 2). In the biological process category, metabolic process (GO:0008152) (young 15391, 60.7% and mature 14395, 62.7%) followed by cellular process (GO:0009987) (young 13691, 54.0% and mature 12381, 53.9%) were prominent in young and mature leaves, suggesting these CDS might be involved in some important metabolic activities in senna. In the molecular function category, “catalytic activity” (GO: 0003824) (young 12555, 49.6% and mature 12049, 52.4%) represented most abundant term, followed by “binding activity” (GO: 0005488) (young 12513, 49.4% and mature 11394, 49.6%). Extremely low percentage of genes were classified in terms of “protein tag” (GO: 0031386), “locomotion” (GO: 0040011), “metallochaperone” (GO: 0016530) and “viral reproduction” (GO: 0016032) in young as well as mature leaf transcriptomes. Under the cellular component category, highest number of CDS was associated with “cell” (GO: 0005623) (young 12790, 50.5% and mature 11377, 49.5%) and “cell part” (GO: 0044464) (young 12790, 50.5% and mature 11377, 49.5%) followed by organelle (GO: 0043226) (young 7382, 29.1% and mature 6580, 28.6%) in young and mature leaf. Both the libraries showed similar type of distribution pattern of CDS under different GO terms.

Bottom Line: A total of 10,763 CDS differentially expressing in both young and mature leaf libraries of which 2,343 (21.7%) CDS were up-regulated in young compared to mature leaf.Several differentially expressed genes found functionally associated with sennoside biosynthesis.We have identified a set of putative genes involved in various secondary metabolite pathways, especially those related to the synthesis of sennosides which will serve as an important platform for public information about gene expression, genomics, and functional genomics in senna.

View Article: PubMed Central - PubMed

Affiliation: ICAR-Directorate of Medicinal and Aromatic Plants Research (DMAPR), Anand, Gujarat, India.

ABSTRACT
Senna (Cassia angustifolia Vahl.) is a world's natural laxative medicinal plant. Laxative properties are due to sennosides (anthraquinone glycosides) natural products. However, little genetic information is available for this species, especially concerning the biosynthetic pathways of sennosides. We present here the transcriptome sequencing of young and mature leaf tissue of Cassia angustifolia using Illumina MiSeq platform that resulted in a total of 6.34 Gb of raw nucleotide sequence. The sequence assembly resulted in 42230 and 37174 transcripts with an average length of 1119 bp and 1467 bp for young and mature leaf, respectively. The transcripts were annotated using NCBI BLAST with 'green plant database (txid 33090)', Swiss Prot, Kyoto Encylcopedia of Genes & Genomes (KEGG), Cluster of Orthologous Gene (COG) and Gene Ontology (GO). Out of the total transcripts, 40138 (95.0%) and 36349 (97.7%) from young and mature leaf, respectively, were annotated by BLASTX against green plant database of NCBI. We used InterProscan to see protein similarity at domain level, a total of 34031 (young leaf) and 32077 (mature leaf) transcripts were annotated against the Pfam domains. All transcripts from young and mature leaf were assigned to 191 KEGG pathways. There were 166 and 159 CDS, respectively, from young and mature leaf involved in metabolism of terpenoids and polyketides. Many CDS encoding enzymes leading to biosynthesis of sennosides were identified. A total of 10,763 CDS differentially expressing in both young and mature leaf libraries of which 2,343 (21.7%) CDS were up-regulated in young compared to mature leaf. Several differentially expressed genes found functionally associated with sennoside biosynthesis. CDS encoding for many CYPs and TF families were identified having probable roles in metabolism of primary as well as secondary metabolites. We developed SSR markers for molecular breeding of senna. We have identified a set of putative genes involved in various secondary metabolite pathways, especially those related to the synthesis of sennosides which will serve as an important platform for public information about gene expression, genomics, and functional genomics in senna.

No MeSH data available.