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2018 - 201912020 - 20231
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Subject: Biochemistry × Author: Heena Rani × Start date: 2001 × Type: Thesis ×
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    Malt quality and thermostability studies in mutants, hulled, hull-less and wild genotypes of barley (Hordeum vulgare L.)
    (Punjab Agricultural University, 2023) Heena Rani; Bhardwaj, Rachana D
    Despite various attempts to develop malt hydrolytic enzymes, the catalytic activity and thermostability of these enzymes are still far from sufficiency for their use in the malting process. The present study explored diverse barley germplasm to identify the genotypes with high activity and thermostability of enzymes alongwith the proteins responsible for high malt yield. Based on enzymatic assays, we selected BL1714T, DWRB137T and BL2105T with highly activity and thermostability of α and β amylases and limit dextrinase, which are key diastatic power enzymes. The barley proteome of these genotypes was analyzed before and after kilning using label-free quantitative global proteomic approach against the control genotypes; BL1718Ctrl, DWRUB64Ctrl and BL2083Ctrl, respectively. Among various isoforms of hydrolytic enzymes Bamy 1 and Amy1_2 were more abundant in thermostable genotypes indicating their role in controlling the activity of these enzymes at high temperatures. Gene ontology enrichment analysis indicated disulfide reductase, exopeptidase, serine-type peptidase (serpin), cell wall hydrolase and oxidoreductase activities as most common enriched terms in thermostable genotypes. These proteins work to release the bound form of hydrolytic enzymes, resulting in their increased overall activity. These proteins could be employed as biochemical indicators for identifying barley lines with high activity and thermostability. Using GC-MS/MS, we identified maltose, glucose, talose, sucrose, fructose, turanose and other minor sugars in green and kilned malt. Talose and turanose were identified among major sugars for the first time during malting. Further, malt quality analysis suggested that βglucanase and malt β-glucan can be used as additional traits to produce malt with superior quality.
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    Expression patterns of glucosinolate transporter genes GTR1 and GTR2 in relation to variation patterns for leaf and seed glucosinolates in Brassica juncea
    (Punjab Agricultural University, Ludhiana, 2018) Heena Rani; Sharma, Sanjula
    The present study was aimed at relating expression patterns of glucosinolates transporter genes (GTR1 and GTR2) with varied pattern of glucosinolate (GLS) content at different developmental stages in mustard (B. juncea). A total of two hundred genotypes were screened for their leaf and seed GLS content and were categorized into two sets. One set with GLS genotypes containing high GLS content (3.29 to 23.18 μmoles/g DW in leaves and 68.17 to 116.38 μmoles/g defatted meal in seeds), and the another set with genotypes containing low GLS content (2.42 to 8.64 μmoles/g in leaves and 24.42 to 30.14 μmoles/g defatted meal in seeds). A huge difference was observed between leaves and seed GLS content indicating the involvement of some other plant tissue as major site for GLS biosynthesis and the information was uncovered by analyzing GLS content in tissues from various developmental stages for GLS content. GLS content increased from open buds (22.02-24.91 μmoles/g DW in high GLS set; 11.17-19.06 μmoles/g DW in low GLS set) through wilted flowers (23.8-35.72 μmoles/g DW in high GLS set; 16.3-21.3 μmoles/g DW in low GLS set) to premature siliques at 14 DAF (43.54-69.14 μmoles/g DW in high GLS set and 21.9-27.68 μmoles/g DW in low GLS set). However, followed by a decrease from 14 DAF through 21 DAF siliques (25.28-42.69 μmoles/g DW in high GLS set; 17.63-25.09 μmoles/g DW in low GLS set) to 30 DAF siliques (9.59-32.68 μmoles/g DW in high GLS set; 9.01-14.7 μmoles/g DW in low GLS set) which indicates the transport of GLS occurs from silique walls towards developing seeds. The results were confirmed by the qRT-PCR gene expression studies where fold change in gene expression of GTR1 and GTR2 increased from open buds (almost negligible) to siliques at 30 DAF (1.93-12.81 fold for GTR1 and 0.38-3.60 fold for GTR2) which indicate an increase in GLS transport as a plant progresses towards maturity. GLS profiling results showed sinigrin as the most abundant GLS in leaves, whereas, gluconapin was highest in seeds. Removal of these anti-nutritional compounds (GLSs) will improve the mustard meal quality and allow its usage as food and feed purpose that could help in eradicating malnourishment problem from the country. Our study could prove helpful in lowering the GLS content by identifying alternate alleles that prevent accumulation of GLSs in developing seeds.