PhD

Ginger (Zingiber officinale Rosc.) is one of the oldest known spices and is much valued for its medicinal properties. Ginger rhizome is a source of active biological compounds which are responsible for its medicinal properties. Gingerol is the major pungent polyphenol in ginger which has got a wide array of pharmacological properties. Research on somaclonal variation done at Kerala Agricultural University could locate ginger somaclones with high gingerol content and in the course of investigations high somaclonal variation was observed for quality components. Ginger has a very big genome of 23,618 Mbp which is little exploited and reports on the genomic base of gingerol biosynthesis are scanty. The present investigations hence aim at profiling the metabolites in selected ginger somaclones using high throughput analytical platforms and to analyze the gene expression with respect to gingerol production. One released ginger variety from KAU (Athira), two selected ginger somaclones (B3 and 132M) and parent cultivar (Maran) formed the experimental materials for the study. Studies were carried out at Centre for Plant Biotechnology and Molecular Biology, Distributed Information Centre of College of Horticulture and Arjuna Natural Extracts Pvt. Ltd., Aluva during August 2013 to July 2017. The profiling of aroma principles using Gas Chromatography-Mass Spectrometry (GC-MS) and pungency principles using High Performance Liquid Chromatography (HPLC) at various growth stages viz. five months after planting (5MAP), six months after planting (6MAP) and seven months after planting (7MAP), revealed that aroma and pungency principles accumulated in ginger rhizomes at the rhizome formation stage (5MAP). Clone to clone variation was observed in the number and quantity of aroma and pungency principles accumulated in the rhizome. Total gingerol content in somaclone B3 (19.07%) was high when compared to the control cultivar Maran (17.49%) irrespective of the growth stages.
Gene expression for Chalcone synthase in selected somaclones done using real time PCR assay showed highest gene expression in somaclone B3 when the control cultivar Maran was set as calibrator. Somaclone B3 recorded 54 per cent increase in Chalcone synthase gene expression over the control cultivar Maran. Suppression subtractive hybridization done to identify differentially expressed genes in somaclone B3 and control cultivar Maran could prepare Expressed Sequence Tag (EST) libraries both for rhizome and leaf. Analysis of EST sequences (25 rhizome ESTs and 19 leaf ESTs) using various bioinformatic tools revealed that there were no differentially expressed genes for gingerol production in rhizome ESTs. But eleven differentially expressed proteins involved in signaling response, protein trafficking, photosynthesis, ATP formation and transposon mediated mutation were observed in rhizome ESTs. The analysis of leaf ESTs showed differential gene expression in somaclone B3 for 3-ketoacyl CoA thiolase (ACAA1) gene which is involved in gingerol biosynthetic pathway. Hence the higher expression of 3-ketoacyl CoA thiolase gene is responsible for the high gingerol content in somaclone B3 as compared to control cultivar Maran. Eighteen other differentially expressed proteins involved in biological processes like transportation of plant secondary metabolites and their intermediates, mobilization of sucrose into pathways involved in metabolism, lipid biosynthesis, transportation of cellular material to microtubules, biogenesis of metabolic pathways in Calvin cycle were observed in leaf ESTs. The differentially expressed gene (ACAA1) can be further validated using northern blotting and quantitative real time PCR by designing specific primers from the ESTs. Expressed sequence tags and corresponding differentially expressed proteins can be used as molecular markers. Post translational modification in differentially expressed proteins can be used to study the mechanism of gingerol production. Forty four sequences deposited at NCBI form the base sequences for further research.