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2015 - 20163
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Subject: Agricultural Biotechnology × End date: 2016 × Start date: 2010 × Type: Thesis ×
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    ThesisItemOpen Access
    Characterization and Identification of Potential Probiotic Lactic Acid Bacteria (LAB) Isolated from Fermented Dairy Product of Assam (Doi)
    (AAU, Jorhat, 2016-07) FATMA, JAIBA; Barooah, M.
    Probiotics are the health promoting viable microorganisms that exhibit a beneficial effect on the health of human being by improving the intestinal microbial balance. Lactic acid bacteria (LAB) are important and well known agents. Fermented milk products like Dahi are known source of probiotic organisms. In the present study, LAB was isolated from 25 different ‘Doi’ samples collected from 8 different districts of Assam viz, Tinsukia, Dibrugarh, Sivsagar, Jorhat, Lakhimpur, Udalguri, Morigaon and Bongaigaon. A total of 30 bacteria were isolated initially but only 20 bacteria isolated from Doi meet the classification of LAB as Gram-positive and catalase negative (Salminen et al., 1993). These 20 LAB isolates were characterized based on their phenotypic, biochemical and molecular characteristics. Further, the isolates were screened for their potential probiotic properties based on conditions simulating human gastrointestinal tract (GI). These isolates were able to survive at low pH 2.5 which is similar to the pH of the human GI tract and able to convert lactose into lactic acid. Further studies revealed that these isolates were able to grow in a temperature ranging from 15-40ºC and grow at 9 % NaCl concentration. The antimicrobial activity of these isolates were tested against six pathogenic bacteria viz, Escherichia coli, Pseudomonas putida, Staphylococcus aureus, Staphylococcus epidermidis, Salmonella enterica and Bacillus cereus. Four isolates (ABT-Z2, ABT-Z3, ABT-Z4 and ABT-Z22) were able to inhibit the growth of all the tested six pathogenic bacteria. All the isolates produced Exopolysaccharide (EPS) whose production was maximum at pH-6. Seven best LAB isolates were characterized by 16S rRNA gene sequencing and sequence analysis using nBLAST revealed that five of the seven isolates belonged to Lactobacillus plantarum and two belonged to L. brevis. The results of the present analysis revealed that strain L. plantarum is the predominant species of LAB found in Assamese Doi. The strain also displayed the best probiotic potential among the tested isolates. In future, this strain of bacteria can be used in preparing probiotic milk based fermented product and can be developed as an efficient starter culture to make Doi.
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    ThesisItemOpen Access
    STUDY OF DIFFERENTIAL GENE EXPRESSION IN RICE UNDER DROUGHT STRESS
    (AAU, Jorhat, 2015-07) Deka, Diganta; Modi, M. K.
    Rice is a dietary staple for a large part of the world’s human population which is grown under varying water regimes ranging from flooded to rainfed upland condition. The predominantly rice-growing areas in Asia are often threatened by severe abiotic stresses, the most common being drought which affects the yield potential of rice across all agro-climatic regions of the globe. Interestingly, some of the rice varieties of Northeast India are found to be drought tolerant e.g. Banglami. Comparative biochemical and physiological analyses of the variety with a high-yielding variety Luit confirmed this. Keeping all these points in view, high throughput RNA-Seq of the variety Banglami in presence and absence of drought was performed as an attempt to study the differential gene expression in Banglami. The RNA isolated by Trizol reagent (Invitrogen) was used for preparation of paired-end libraries using Illumina TruSeq RNA Library Preparation Kit. Libraries were sequenced using 2 X 150 PE chemistry on NextSeq. The reads were aligned against the indica reference assembly (ASM465v1). The expression analysis of the genes revealed 25,272 and 24,408 numbers of expressed genes in well watered control and drought stressed sample, respectively. Further analysis revealed 391 numbers of genes showing differential expression among which 86 were up-regulated and 305 were down-regulated. Among the differential expressed genes a number of genes were found to be very important for development of drought stress tolerant behaviour. Further, downstream analysis like Gene Ontology enrichment analysis, KEGG pathway analysis and QTL mapping were also performed which revealed important informations regarding the differentially expressed genes under drought stress condition in particular and the whole transcriptome of the Banglami variety of rice in general. The present study identified altered gene expression in rice induced by drought stress and provided a comprehensive map of drought responsive genes and pathways. Thus the results of the present investigation can serve as valuable genetic resource for gene expression, genomics and functional genomics studies in general and drought stress research in rice in particular.
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    ThesisItemOpen Access
    Transcriptome sequencing of Cymbopogon winterianus and characterization of metabolic pathway genes
    (AAU, Jorhat, 2015) Devi, Kamalakshi; Sen, Priyabrata
    Citronella (Cymbopogon winterianus) is an aromatic and medicinal grass grown for commercial and industrial purpose due to its large repository of isoprenoids compounds known for its anti-tumoral, antibacterial, anti-fungal, antiviral, anti-cancerous and detoxifying properties as well as use as natural insect repellent properties. Despite of having numbers of valuable utilities, the genes involved in the biosynthetic pathway of these terpenes in citronella are not yet clearly elucidated. Keeping these points in view, the present study is formulated as a pioneering attempt to generate a large amount of molecular level information for citronella followed by identification of secondary metabolite associated gene. The whole transcriptome sequencing of leaf and root tissues of citronella (Cv. Jorlab-1) was carried out on Illumina MiSeq NGS plateform using Paired end (PE) 2x250bp library. A total of 11.7 Gb data were generated which upon de novo assembly yielded 68,986 unigenes. The complete bioinformatic analysis of these genes resulted : functional annotation of 63,454 unigenes along with 5478 novel genes; GO annotation of 56,621 unigenes; GO enrichment analysis of 218 secondary metabolite associated genes; detection of 3,851 putative SSR motifs and KEGG pathway analysis annotated 3,362 unigenes. The RT and qRT PCR analysis of the 20 secondary metabolite associated genes were in line with the findings from differential expression study. The regulatory relation of miRNA & TF with secondary metabolite associated genes revealed 14 pre-miRNA targets 29 TFs and 4 genes. There were three key regulatory enzymes of terpenoid backbone synthesis pathway viz.1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR) (GenBank: KJ749651), 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMGR) (GenBank: KM504513) and Geranylgeranyl pyrophosphate synthase (GGPPS) (GenBank: KP017769) of citronella analyzed further. The full length gene sequence of these genes were obtained by performing 5’ RACE PCR. The complete in silico proteomics analysis of the three proteins included analysis of primary protein sequence, phylogeny, physico-chemical property of the proteins, homology modeling and model validation along with molecular dynamic simulation and finally docking of co factor (NADPH). The 3D structure analysis of the proteins gave a clear insight about the catalytic sites, substrate and cofactor binding pockets and various conserved motifs of these proteins. It can be concluded that, this work sets the stage for multi-faceted future improvement of the plant, through discovery of new genes, marker-assisted breeding or genetic engineering, on this species as well as for other species of Poaceae and terpene producing medicinal plants.