Molecular cloning and differential expression analysis of a squalene synthase gene from Dioscorea zingiberensis, an important pharmaceutical plant

Ye, Yun; Wang, Runfa; Jin, Liang; Shen, Junhao; Li, Xiaotong; Yang, Ting; Zhou, Mengzhuo; Zhi-fan, Yang; Chen, Yongqin

doi:10.1007/s11033-014-3487-9

Cited by 23 publications

(10 citation statements)

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“…Squalene synthase (SQS) catalyzes the condensation of two farnesyl pyrophosphate (C15 unit) to 30 carbons compound squalene, the first committed intermediate in sterol and triterpene saponin biosynthesis pathway [ 56 ]. Squalene synthase is well characterized in various plants like Chlorophytum borivilianum, [ 57 ] Dioscorea zingiberensis, [ 58 ] and Siraitia grosvenorii [ 56 ]. Squalene epoxidase (SQE) with the cofactors O 2 and NADPH catalyzes the conversion of squalene to 2,3-oxidosqualene that acts as a substrate for various oxidosqualene cyclases [ 59 ].…”

Section: Discussionmentioning

confidence: 99%

De novo sequencing, assembly and characterisation of Aloe vera transcriptome and analysis of expression profiles of genes related to saponin and anthraquinone metabolism

et al. 2018

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BackgroundAloe vera is a perennial, succulent, drought-resistant plant that exhibits many pharmacological characteristics such as wound healing ability against skin burns, anti-ulcer, anti-inflammatory, anti-tumor, anti-viral, anti-hypercholesterolemic, anti-hyperglycemic, anti-asthmatic and much more. Despite great medicinal worth, little genomic information is available on Aloe vera. This study is an initiative to explore the full-scale functional genomics of Aloe vera by generating whole transcriptome sequence database, using Illumina HiSeq technology and its progressive annotation specifically with respect to the metabolic specificity of the plant.ResultsTranscriptome sequencing of root and leaf tissue of Aloe vera was performed using Illumina paired-end sequencing technology. De novo assembly of high quality paired-end reads, resulted into 1,61,733 and 2,21,792 transcripts with mean length of 709 and 714 nucleotides for root and leaf respectively. The non-redundant transcripts were clustered using CD-HIT-EST, yielding a total of 1,13,063 and 1,41,310 unigenes for root and leaf respectively. A total of 6114 and 6527 CDS for root and leaf tissue were enriched into 24 different biological pathway categories using KEGG pathway database. DGE profile prepared by calculating FPKM values was analyzed for differential expression of specific gene encoding enzymes involved in secondary metabolite biosynthesis. Sixteen putative genes related to saponin, lignin, anthraquinone, and carotenoid biosynthesis were selected for quantitative expression by real-time PCR. DGE as well as qRT PCR expression analysis represented up-regulation of secondary metabolic genes in root as compared to leaf. Furthermore maximum number of genes was found to be up-regulated after the induction of methyl jasmonate, which stipulates the association of secondary metabolite synthesis with the plant’s defense mechanism during stress. Various transcription factors including bHLH, NAC, MYB were identified by searching predicted CDS against PlantTFdb.ConclusionsThis is the first transcriptome database of Aloe vera and can be potentially utilized to characterize the genes involved in the biosynthesis of important secondary metabolites, metabolic regulation, signal transduction mechanism, understanding function of a particular gene in the biology and physiology of plant of this species as well as other species of Aloe genus.Electronic supplementary materialThe online version of this article (10.1186/s12864-018-4819-2) contains supplementary material, which is available to authorized users.

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Section: Discussionmentioning

confidence: 99%

De novo sequencing, assembly and characterisation of Aloe vera transcriptome and analysis of expression profiles of genes related to saponin and anthraquinone metabolism

et al. 2018

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“…The activity of squalene synthase formation was assayed as follows (Ye et al, 2014). The 500 μL reaction mixture contained 100 mM Tris–HCl (pH 7.5), 10 mM farnesyl pyrophosphate triammonium salt (FPP; Sigma–Aldrich, USA), 10 mM MgCl 2 , 1 mM DTT, 2% Glycine, 3 mM NADPHNa 4 and 475 μL crude enzyme solution prepared from the E. coli cells.…”

Section: Methodsmentioning

confidence: 99%

Molecular Cloning and Functional Analysis of Squalene Synthase 2(SQS2) in Salvia miltiorrhiza Bunge

et al. 2016

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Salvia miltiorrhiza Bunge, which is also known as a traditional Chinese herbal medicine, is widely studied for its ability to accumulate the diterpene quinone Tanshinones. In addition to producing a variety of diterpene quinone, S. miltiorrhiza Bunge also accumulates sterol, brassinosteroid and triterpenoids. During their biosynthesis, squalene synthase (SQS, EC 2.5.1.21) converts two molecules of the hydrophilic substrate farnesyl diphosphate (FPP) into a hydrophobic product, squalene. In the present study, cloning and characterization of S. miltiorrhiza Bunge squalene synthase 2 (SmSQS2, Genbank Accession Number: KM408605) cDNA was investigated subsequently followed by its recombinant expression and preliminary enzyme activity. The full-length cDNA of SmSQS2 was 1 597 bp in length, with an open reading frame of 1 245 bp encoding 414 amino acids. The deduced amino acid sequence of SmSQS2 shared high similarity with those of SQSs from other plants. To obtain soluble recombinant enzymes, the truncated SmSQS2 in which 28 amino acids were deleted from the carboxy terminus was expressed as GST-Tag fusion protein in Escherichia coli BL21 (DE3) and confirmed by SDS-PAGE and Western Blot analysis, and the resultant bacterial crude extract was incubated with FPP and NADPH. Gas chromatograph-mass spectrometer analysis showed that squalene was detected in the in vitro reaction mixture. The gene expression level was analyzed through Quantitative real-time PCR, and was found to be higher in roots as compared to the leaves, and was up-regulated upon YE+ Ag+ treatment. These results could serve as an important to understand the function of the SQS family. In addition, the identification of SmSQS2 is important for further studies of terpenoid and sterol biosynthesis in S. miltiorrhiza Bunge.

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“…Two other specific primers, Actin-qF and Actin-qR, were designed to amplify the housekeeping gene β-actin as an internal control (Ye et al 2014). For tissue expression profiles, total RNA was extracted from young leaves, medium leaves, mature leaves, old leaves, young stems, old stems, young rhizomes, and old rhizomes of D. zingiberensis using the Universal Plant Total RNA Extraction Kit (Bioteke), respectively.…”

Section: Dzs3gt Expression Dzs3gt Activity and The Accumulation Of Smentioning

confidence: 99%

Identification and functional characterization of DzS3GT, a cytoplasmic glycosyltransferase catalyzing biosynthesis of diosgenin 3-O-glucoside in Dioscorea zingiberensis

Song

Zhang

et al. 2017

Plant Cell Tiss Organ Cult

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Molecular cloning and differential expression analysis of a squalene synthase gene from Dioscorea zingiberensis, an important pharmaceutical plant

Cited by 23 publications

References 43 publications

De novo sequencing, assembly and characterisation of Aloe vera transcriptome and analysis of expression profiles of genes related to saponin and anthraquinone metabolism

De novo sequencing, assembly and characterisation of Aloe vera transcriptome and analysis of expression profiles of genes related to saponin and anthraquinone metabolism

Molecular Cloning and Functional Analysis of Squalene Synthase 2(SQS2) in Salvia miltiorrhiza Bunge

Identification and functional characterization of DzS3GT, a cytoplasmic glycosyltransferase catalyzing biosynthesis of diosgenin 3-O-glucoside in Dioscorea zingiberensis

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