Squalene synthase (SQS) is a rate-limiting enzyme necessary to produce pentacyclic triterpenes in plants. It is an important enzyme producing squalene molecules required to run steroidal and triterpenoid biosynthesis pathways working in competitive inhibition mode. Reports are available on information pertaining to SQS gene in several plants, but detailed information on SQS gene in Gymnema sylvestre R. Br. is not available. G. sylvestre is a priceless rare vine of central eco-region known for its medicinally important triterpenoids. Our work aims to characterize the GS-SQS gene in this high-value medicinal plant. Coding DNA sequences (CDS) with 1245 bp length representing GS-SQS gene predicted from transcriptome data in G. sylvestre was used for further characterization. The SWISS protein structure modeled for the GS-SQS amino acid sequence data had MolProbity Score of 1.44 and the Clash Score 3.86. The quality estimates and statistical score of Ramachandran plots analysis indicated that the homology model was reliable. For full-length amplification of the gene, primers designed from flanking regions of CDS encoding GS-SQS were used to get amplification against genomic DNA as template which resulted in approximately 6.2-kb sized single-band product. The sequencing of this product through NGS was carried out generating 2.32 Gb data and 3347 number of scaffolds with N50 value of 457 bp. These scaffolds were compared to identify similarity with other SQS genes as well as the GS-SQSs of the transcriptome. Scaffold_3347 representing the GS-SQS gene harbored two introns of 101 and 164 bp size. Both these intronic regions were validated by primers designed from adjoining outside regions of the introns on the scaffold representing GS-SQS gene. The amplification took place when the template was genomic DNA and failed when the template was cDNA confirmed the presence of two introns in GS-SQS gene in Gymnema sylvestre R. Br. This study shows GS-SQS gene was very closely related to Coffea arabica and Gardenia jasminoides and this gene harbored two introns of 101 and 164 bp size.

中文翻译：

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匙羹藤角鲨烯合酶基因的表征。

角鲨烯合酶 (SQS) 是植物中产生五环三萜所必需的限速酶。它是一种重要的酶，可产生角鲨烯分子，以竞争性抑制模式运行甾体和三萜类生物合成途径。有关于几种植物中 SQS 基因信息的报告，但有关于匙羹藤 SQS 基因的详细信息。不可用。G. sylvestre 是中央生态区的一种无价的稀有藤本植物，以其具有重要药用价值的三萜类化合物而闻名。我们的工作旨在表征这种高价值药用植物中的 GS-SQS 基因。长度为 1245 bp 的编码 DNA 序列 (CDS) 代表从 G. sylvestre 中的转录组数据预测的 GS-SQS 基因，用于进一步表征。为 GS-SQS 氨基酸序列数据建模的 SWISS 蛋白质结构的 MolProbity 得分为 1.44，Clash 得分为 3.86。拉马钱德兰图分析的质量估计和统计得分表明同源模型是可靠的。对于基因的全长扩增，使用从编码 GS-SQS 的 CDS 的侧翼区域设计的引物以针对作为模板的基因组 DNA 进行扩增，从而产生大约 6.2-kb 大小的单带产物。该产品通过 NGS 进行测序，产生 2.32 Gb 数据和 3347 个支架，N50 值为 457 bp。比较这些支架以鉴定与其他 SQS 基因以及转录组的 GS-SQS 的相似性。代表 GS-SQS 基因的 Scaffold_3347 含有两个 101 和 164 bp 大小的内含子。这两个内含子区域均通过从代表 GS-SQS 基因的支架上内含子的相邻外部区域设计的引物进行验证。当模板为基因组 DNA 时发生扩增，当模板为 cDNA 时扩增失败，证实了匙羹藤中 GS-SQS 基因中存在两个内含子。该研究表明 GS-SQS 基因与小粒咖啡和栀子花非常密切相关，该基因包含两个大小分别为 101 和 164 bp 的内含子。