Sen, PriyabrataSaha, Oliva2022-02-262022-02-262018-07https://krishikosh.egranth.ac.in/handle/1/5810182944Citronella (Cymbopogon winterianus), an aromatic and medicinal grass from Poaceae family, is grown for the commercial and industrial purpose. It has the large repository of isoprenoid compounds known for its anti-tumoral, anti-bacterial, anti-fungal, anti-viral, anticancerous, detoxifying and natural insect repellent properties. Out of the different components of citronella oil, linalool (3,7-Dimethylocta-1,6-dien-3-ol) is one of the significant aromatic ingredient. Formation of linalool is catalysed by linalool synthase (LIS) which is encoded by linalool synthase gene. This gene has been widely studied in many medicinal plants along with sorghum, rice, maize, setaria, etc.However, till date, no studies on molecular cloning, characterization and tissue-specific expression profiling of LIS gene from C. winterianus (CwLIS) has been reported.Therefore, the present study aims at cloning and characterization of full-length cDNA of LIS gene from C. winterianus. The full-length sequence of CwLIS was obtained by primer walking using several pairs of degenerate primers and the sequence fragments were aligned to acquire full length sequence. Cloning of the desired gene (1758bp) was performed using TOPO TA vector (3.9kb), and the transformation was done in E. coli DH5α competent cells. The sequence analysis revealed 1758 bps cDNA, which has a Coding Sequence (CDS) of 1422 bps that encodes a protein of 473 amino acids. The domain analysis revealed that CwLIS is a single domain protein under terpene synthase superfamily. The multiple sequence alignment (MSA) based on PSI-BLAST search against RefSeq with 62% sequence identity revealed the presence of two aspartate-rich regions, which are supposed to coordinate 3 Mg2+ ions. The phylogenetic analysis revealed that the CwLIS is closely related to Sorghum bicolor linalool synthase. To understand the molecular mechanism behind Mg2+ and linalool binding, first, the 3D model of CwLIS was generated and validated with their stereo chemical parameters. Further, to find out the expression profile of CwLIS in different tissues, Reverse Transcriptase-PCR was performed and the results revealed a higher expression in leaf sheath followed by leaf, root and flower and further conformed by qRTPCR analysis. The result from the present study provide basic information for further research about linalool synthase and comprehensive sequence resource for study, such as gene expression, genomics and functional genomics in Cymbopogon winterianus.EnglishThesis