Theses

Browse

Search Results

Now showing 1 - 9 of 17

Open Access
GENE EXPRESSION STUDIES FOR LOW TEMPERATURE TOLERANCE AT THE BOOTING STAGE IN RICE
(2023) Bora, Smriti Shyamolee; Baruah, Akhil Ranjan
Rice (Oryza sativa L.) is a staple food crop for half of the world’s population and it has been grown as major cereal crop in three distinct seasons in Assam. Out of various constrains of rice production, abiotic stresses such as drought, low temperature (LT), salinity adversely affect crop growth and development. Low temperature stress at the early growth and reproductive stages in rice are critical, and the present study was conducted to screen low temperature tolerant genotypes at the booting stage (intermediate stage of reproductive phase) and study gene expression for cold responsive genes in tolerant and susceptible rice genotypes. A total of 30 rice cultivars collected from Regional Agricultural Research Station, Karimganj, AAU were subjected to low temperature stress at the booting stage in a growth chamber maintained at 12- 12.5C for seven days. The recovery and low temperature tolerance of plants after stress were evaluated based on pollen fertility, number of panicle emerged, number of seeds set and grain yield. The results revealed that the phenotypes of C-59 and IET-9097 were superior for all the traits under study whereas Costco and IET-8687 showed lowest trait values, indicating varied levels of low temperature tolerance at the booting stage. These four cultivars, C-59 and IET- 9097 as low temperature tolerant; Costco and IET-8687 as low temperature susceptible were chosen to study gene expression analysis using six cold responsive genes (OsCOLD1, OsDREB2A, OsCTB1, OsLTT1, OsAPX1, OsNAC9) through real time quantitative reverse transcriptase polymerase chain reaction (RT-qPCR). The results showed that all the genes were up-regulated in the cold tolerant cultivars, C-59 and IET-9097 and down regulation in the other two cold susceptible cultivars, Costco and IET-8687. After performing qRT-PCR analysis, correlation between the gene expression data and phenotypic observations followed by path analysis was performed to detect associations between the genes and phenotypes. Path analysis revealed that out of six genes used in the study, three genes showed direct positive effect over grain yield (OsDREB2A, OsCTB1 and OsNAC9) and three genes showed indirect effect for grain yield (OsCOLD1, OsLTT1 and OsAPX1). The study is preliminary with respect to low temperature tolerance at the booting stage using rice collection of AAU and hence, further validation is required.
Open Access
STUDY ON GENETIC FIDELITY IN TISSUE CULTURED RAISED PLANTLETS OF CITRUS LIMON (ASSAM LEMON)
(2023) Das, Ratnakishore; Sen, Priyabrata
Citrus lemon, a highly significant species both economically and nutritionally, is extensively cultivated in Assam. The demand for this valuable crop has led to the development of efficient methods for large-scale multiplication to meet the increasing requirements of the market. Micropropagation, an advanced tissue culture technique, offers a promising approach for the rapid production of genetically identical plants. In this study, Citrus lemon was micropropagated to assess its potential for large-scale multiplication, and the genetic fidelity of the micropropagated plantlets was evaluated using Simple Sequence Repeat (SSR), Inter-Simple Sequence Repeat (ISSR), and long Retrotransposon (LTR) markers. SSR markers are highly polymorphic, consisting of short repetitive DNA sequences, while ISSR markers target the regions between microsatellites. LTR markers, on the other hand, focus on retrotransposon dispersed throughout the genome Healthy and vigorously growing shoot-tip were excised from donor plants and placed on the medium. Within a few weeks, these shoots successfully developed into multiple plantlets through the process of organogenesis. The micropropagated plantlets were then acclimatized to the greenhouse environment, and their growth performance was monitored. To determine the genetic stability and fidelity of the micropropagated plantlets. Fifty plant samples including the mother plant and rest in vitro regenerated plantlets are used for testing the genetic fidelity. Total genomic DNA was extracted from both the mother plants and the micropropagated plantlets using standard protocols, and the SSR, ISSR, and LTR markers were employed to assess genetic variation, if any, between the two sets of samples. The results of the genetic fidelity analysis demonstrated no detectable variation between the micropropagated Citrus lemon plantlets and their mother plants. All the SSR, ISSR, and LTR markers revealed identical banding patterns, indicating the absence of any somaclonal variation during the micropropagation process. The amplified fragments of the micropropagated plantlets precisely matched those of the donor plants, confirming the maintenance of genetic uniformity in the micropropagation protocol. In conclusion, this study demonstrates that, successful micropropagation of Citrus lemon with retained genetic fidelity is of immense significance for large-scale multiplication and commercial production.
Open Access
MORPHOLOGICAL AND MOLECULAR CHARACTERIZATION OF VESICULAR ARBUSCULAR MYCORRHIZA ASSOCIATED WITH LEGUMES
(2023) Kudve, Rajkumar Laxman; Nath, Tankeswar
Plant health and soil fertility are largely influenced by nos. of associations between plant and microbes. Among these, the association between Vesicular Arbuscular Mycorrhiza (VAM) and the plant is well-known. VAM fungi create symbiotic association with the roots of plants and increase plant growth largely through phosphorus nutrition. The International Collection of Vesicular Arbuscular Mycorrhiza (INVAM), University of Kansas, U.S. reported that around 160 species of Arbuscular Mycorrhizal Fungi (AMF) have been described by spore morphology. However, in nature there may be great variation of spore‟s morphology within a VAM species. Hence, it is necessary to conduct research on VAM to analyze genus diversity in the soil and for their use as biofertilizer. However, VAM have not been studied extensively at morphological and molecular level by regional diversity like other beneficial microorganisms. Again, the molecular level identification is necessary due to most of the mycorrhizal species are structurally similar, and hence they cannot be differentiated on the basis of their morphological observations. Therefore, in this present study an attempt had been made to isolate and characterize VAM indigenous to Assam, at morphological and molecular levels. For this, VAM spore had been isolated from the rhizospheric soil of two legume crops, Dhaincha (Sesbania bispinosa) and Subabul (Leucaena leucocephala) from ICR farm AAU, Jorhat. At first, morphological identification of the isolated VAM fungi was done under microscope. Spore, sporocarps and hyphal structure were observed by mounting with polyvinyl alcohol-lactic acid-glycerol (PVLG). On the basis of morphological observations (spore color, spore shape and hyphal color) they are supposed to come under the genus, Glomus, Gigaspora, Acaulospora and Scutellospora. For molecular characterization, the isolated VAM fungal spores inoculated with maize as the trap plant in pot culture technique. Further, the presence of arbuscule, vesicle and hyphae of VAM in infected roots of maize seedling were studied under microscope after staining with trypan blue. The study was further extended to molecular characterization by using both universal fungal primers [NS1(F) and NS2(R)] as well as mychorrhyza specific primers [AML1 (F) and AML2 (R)]. Finally, the mycorrhizal isolates were identified as Glomus, Gigaspora, Acaulospora and Scutelospora species. Thus, the tentative classification of AMF done on the basis of morphological characteristics was supported by the molecular characterizations. This was a preliminary study of rhizospheric AM fungi associated with legumes. However, this study can further be extended to study the diversity of plant associated mycorrhizal fungi in Assam soils which will help to develop mycorrhiza based biofertilizer specific to this region. Furthermore, it will give a picture of mycorrhizal diversity of Assam, understanding of which is important to maintain productivity and stability of ecosystem of this region.
Open Access
Molecular basis of non -host resistance to Helicoverpa armigera
(2023) Kanth, Deepak Ranjan; Acharjee, Sumita
The complex molecular mechanism underlying resistance in non-host crops against Helicoverpa armigera is yet to be elucidated. Non-host resistance exhibits remarkable efficacy and broad-spectrum capabilities in countering potential pests, hence, has proven to be a valuable resource for understanding the basis of resistance. Our lab previously studied the molecular mechanism of legume (chickpea and pigeon pea) interactions with H. armigera using transcriptomic and proteomics approaches. Several cell receptor-like kinases (RLKs)/receptor-like proteins (RLPs) and defense-related genes were identified in these studies. In this present investigation, an effort has been made to understand the changes in expression patterns of a few selected cell receptor-like kinases (RLKs)/receptor-like proteins (RLPs), and defense-related genes in response to simulated herbivory (Oral Secretions/OS) and exogenous application of hormones such as Jasmonic acid (JA), Salicylic acid (SA) and Ethephon (ET). The generation of Reactive Oxygen Species (ROS) in samples after 12 h of oral secretion (OS) treatment confirmed that various elicitors were present in OS for effective induction of defense in groundnut. Gene expressing analysis showed significant (>5-fold) upregulation of the Proline-rich RLK PERK 15 gene due to the application of OS. Moreover, the expression of defense-related genes such as Endoglucanase 8-like, PR10, ABA-responsive protein ABR17, Galactose-binding lectin, and Cytochrome P450 90B1 also showed upregulation after 24 h of OS treatment. However, the expression of the above genes was downregulated after 48 h of treatment. It was also observed that the expression of lysM domain RLK 3, ABA-responsive ABR17, and PR10 genes was regulated by JA and ET pathways while galactose-binding lectin gene expression was mediated by the SA pathway. The expression of the endoglucanase 8-like gene showed an antagonistic regulation by JA and SA. Moreover, the trypsin inhibition (TI) assay revealed the expression of TI in the treated samples. Thus, the above results suggest that during the interaction H. armigera, elicitors present in OS are perceived by lysM domain RLK3 receptor in a non-host, groundnut, which activated the downstream signaling pathways to upregulate the expression of various defense-related genes. It would be interesting to study the role of galactose-binding lectin in the non-host defense against H. armigera. The present study shed light on the non-host resistance mechanism using the groundnut-Helicoverpa interactions as a model, which revealed changes in the groundnut gene expression pattern due to simulated herbivory.
Open Access
Expression analysis of pathogenesis-related (PR) genes in Brassica rapa upon Alternaria brassicicola infection
(2023) R, Aryasree; Bhorali, Priyadarshini
Alternaria blight caused by Alternaria brassicicola is the most widespread fungal disease of oilseed Brassicas. Conventional breeding to develop Alternaria resistant cultivars has not been successful due to non-availability of suitable resistance sources. For the development of resistant cultivars, identification of important defense or resistance-related genes is needed. Plants activate different biological pathways upon infection by pathogens and a large number of defense-related genes are modulated during such interactions locally as well as systemically. In a previous study, RNAseq of a wild relative of Brassica, Sinapis alba, was carried out in which several defense related genes were found to be highly upregulated during Alternaria infection. Based on the results of this study, the present research was undertaken to investigate and validate the gene expression patterns of selected gene candidates in the susceptible cultivar B. rapa var. Toria (TS-38) in order to check the function of these genes in conferring resistance. To initiate the study, infected tissues of a highly susceptible Toria variety TS-38, showing characteristic symptoms of Alternaria blight were collected for isolation and purification of the pathogen. On the basis of conidial morphology as observed through microscopic studies, the pathogen was identified to be Alternaria brassicicola. Molecular detection of the fungus was carried out by amplification using ITS primers followed by sequencing. Pathogenicity test was done through detached leaf assay and seedling inoculation test and confirmation was done by testing the Koch postulates. After pathogen inoculation, ROS generation and proline accumulation were detected during the biotic stress. Staining with trypan blue helped to visualise necrosis in response to A. brassicicola infection in the leaf tissues. Finally, total RNA was isolated from the B. rapa inoculated (and control) leaf tissues followed by cDNA preparation for real time qRT-PCR analysis of gene expression. Primers available from the previous study, which were designed using S. alba sequences of some important gene candidates, including PR and resistance-related genes were used for the expression analysis. The results depict that all the PR genes were upregulated at lower levels at 24 hpi. Later, although the expression increased towards 48 hpi, it gradually decreased. The resistance related genes also showed upregulation with peak expression at 48 hpi, but later downregulated. Interestingly, one disease resistance (DR) gene showed downregulation across all the time points in the susceptible cultivar and and could be an important gene responsible for resistance. Thus, all the genes are clearly implicated to have a role in active defense responses. As a future prospect, complete functional characterization of such genes could be done as they would serve as important potential candidates for developing disease resistant varieties of Brassica.
Open Access
COMPARATIVE EVALUATION OF ACARICIDAL PROPERTIES OF Datura stramonium and Datura metel
(AAU, Jorhat, 2020) GOSWAMI, MANISHA PRIYAM; Nath, Tankeswar
Ticks are group of arthropod vectors that not only cause disease but also transmit diverse pathogens that affect human and animal health. Indiscriminating use of synthetic acaricides have led to development of acaricide resistance in ticks. Several plant secondary metabolites are known to exhibit toxic affect on insects. Phyto-extracts are also environmentally friendly and have thus become attractive alternative in insect management strategies. Datura has long since been used in folklore medicine and is reported to have other pharmaceutical uses. My research involved the use of two different species of Datura, viz., D. stramonium and D. metel that grow abundantly in the North East India especially, in Assam. The two species of Datura were collected from different locations of Jorhat district and evaluated for their acaricidal properties. Chemo-profiling of both the plant species revealed the presence of some phyto-metabolites with known biocidal properties, more specifically acaricidal properties. Saponin, Flavonoids, Tannins, Phlobatannins and Steroids were analyzed in both the species and revealed that except for Saponin all others metabolites were present in both the species of Datura. Atropine and Scopolamine earlier reported for their acaricidal activity were assayed using HPTLC technique. Shoots of D. stramonium showed the presence of Atropine (0.04238%) and Scopolamine (0.21554%) whereas; the presence of Scopolamine (0.00754%) was detected only in the roots. Presence of Atropine (0.08083%) only was detected in shoots of D. metel. The variation in Atropine as well as Scopolamine in roots and shoots of the same plant species might be due to variance in tissue specific concerned gene(s) expression, and among the different species might be due to genetic variation or variation in eco-geographical conditions which are yet to be explored. Acaricidal properties of both the plant species were assayed in terms of LC50 as well as Inhibition in Oviposition (IO%) against a tick line of Rhipicephalus microplus collected from Assam. Between the two plant species, D. metel showed lower LC50 (6.37%) than D. stramonium (12.27%). In case of IO also, more IO value (76%) was observed in D. metel than D. stramonium (35.27%). The lower LC50 and higher IO values indicate that D. metel has more acaricidal potential than D. stramonium. Findings from the study also showed that shoots of D. stramonium contains both Atropine (0.04238%) and Scopolamine (0.21554%) whereas, D. metel-shoots contain only Atropine (0.08083%) and it is almost double of what D. stramonium contains. The present study indicates that Atropine may be directly associated with acaricidal activity with little or no role of Scopolamine. Although, the study did not involve the analysis of other compounds, their involvement in acaricidal activity cannot be denied. This research can be extended further to explore specific metabolites having acaricidal properties present in other plant systems including Datura for the better and greater interest of development of plant based-based acaricides/insecticides.
Open Access
Molecular characterization of the gut microbes of greater wax moth (Galleria mellonella)
(AAU, Jorhat, 2020-01) Saikia, Silpi Shikha; Borah, Basanta Kumar
Greater wax moth (GWM), Galleria mellonella (Lepidoptera: Pyralidae) is a notorious honey-bee pest found throughout the world. The moth is very destructive to the honeycomb, lays eggs and their larva grows up eating beewax. Beewax is a natural polymer, mainly composed of saturated/ unsaturated, linear/ complex monoesters, hydrocarbons etc. The most frequent hydrocarbon bond in beewax is the CH2-CH2 which is found in polyethylene (PE). Meanwhile, biodegradation of PE by gut microbes of Indian meal moth, Plodia interpunctella and lesser wax moth (a close species of GWM) was reported. As wax-digestion is not a common character of animals, we hypothesized to encounter some microbes in GWM-gut, which could also be responsible for the PE degradation. Therefore, we aimed this investigation to isolate and identify the gut-microbes from GWM following culture-dependant approach. We had characterized several bacterial and fungal species based on culture characteristics, Gram-staining and several biochemical tests such as carbohydrate utilization test, catalase test, citrate utilization test, MR-VP test, motility test etc. Using 16S-rDNA sequencing with conserved primer-sets from representative types, thirteen bacteria and one microalgae were obtained from the digestive tract of G. mellonella. These species includes Gram-positive Exiguobacterium aestuarii, Bacillus circulans, Microbacterium zaea, Microbacterium paraoxydans, Enterococcus faecalis and Gram-negative Agrobacterium sp., Sphingomonas pseudosanguinis, Sphingobium yanoikuyae, Acinetobacter radioresistens, as well as a microalgae (Picochlorum oklahomensis). Several of these species/ isolates have been reported to degrade polycyclic aromatic hydrocarbon, low density polyethylene, 2-methylphenanthrene etc. Interestingly, P. oklahomensis is a marine microalgae; it steals genes from bacteria and adapt themselves to abiotic stress. Further investigation will be required to find out more precise details about plastic degrading candidate microbes and their biotechnological applications.
Open Access
VALIDATION OF DROUGHT RESPONSIVE miRNA IN A DROUGHT TOLERANT RICE CULTIVAR
(AAU, Jorhat, 2020-10) Debsarma, Oindrila; Sen, Priyabrata
Drought like situation in non-irrigated agriculture system often limits rice production, necessitating introduction of drought tolerance trait into the cultivar of interest. In Assam, effect of dry spells on varieties cultivated on low lands was reported to be reduced up to 43.07%. miRNAs, a class of abundant small noncoding RNAs, have been identified as important regulators of gene expression in both plants and animals and are involved in many aspects of plant development, including the modulation of plant response to stress. Despite technical hurdles to miRNA functional analysis, there is a growing body of evidence that alteration of miRNA accumulation plays an important role in reprogramming plant responses to biotic and abiotic stresses. Assam having a diverse germplasm of rice, might have evolved an unique response to various stress. ARC-10372, a proven drought tolerant landrace may be a good source of drought related miRNAs and can provide insight into the role of miRNA in drought stress. In a previous effort in the Dept. to identify novel variety specific miRNA, small RNA sequencing had been employed to systematically investigate the tissue specific miRNAs responsible to drought stress, which are understudied in rice. The study revealed few novel miRNA in addition to known under drought stress. Differentially expressed miRNA as revealed by sequencing data are confirmed experimentally under drought situation using qRT-PCR. Out of 31 miRNAs, 7 known and 5 novel miRNAs were found to respond to drought stress. The identified target for these miRNAs revealed several conserved miRNAs targeting transcription factors like homeodomain-leucine zipper, MADS box family protein, zinc finger protein and Myb, well known for their importance in drought tolerance in plants. Drastic decrease in abundance of a few of the members of the novel miRNA ( nmiR4, nmiR25, nmiR32, nmiR84 & nmiR86) might suggests important role of these miRNAs in drought tolerance. Further work on revealing its confirmatory role shall yield more information in this regard.
Open Access
IDENTIFICATION AND VALIDATION OF DROUGHT-RESPONSIVE GENES IN THE UPLAND RICE CULTIVAR ‘BANGLAMI
(AAU, Jorhat, 2020-10) Borah, Akankshita; Modi, M.K.
Drought is one of the most important and highly unpredictable abiotic stresses causing drastic reductions in yield under rainfed rice environments, affecting 20% of the total rice-growing area in Asia (Pandey and Bhandari, 2008). Assam has diverse germplasm of rice which includes Banglami, a traditional drought-tolerant Ahu rice cultivar found locally in the state. Since 2013, crossing has been carried out between this drought-tolerant cultivar and Ranjit, a high yielding but drought-susceptible elite cultivar. Many drought-related QTLs were identified and reported from this cross-population. For the current study, twenty QTLs identified in the F4 population were selected for mapping on the parental genome. The results revealed that 17 of the 20 QTLs were present on the parent genomes where 11 of the 20 QTLs (qEBT3.1, qEBT6.2, qNOT6.1, qNOT2.1, qPL1.1, qPL1.2, qPL9.1, qPL9.2, qPL1.1, qNOG9.1, qNOG12.1) were found common to both the cultivars. From these, two of the QTLs (qGY1.1, qGY7.1) were found to be exclusively contributed by Banglami while four of the QTLs (qRLWC9.1, qRLWC9.2, qPH7.1, qDTF12.2) were found to be contributed by Ranjit to the cross-population. Genes associated with the QTLs were identified from whole-genome sequence data of both the cultivars. In total, 2454 genes were located in the 13 QTLs found in Banglami while 1585 genes were found in Ranjit in the 15 QTLs. The differential expression of the candidate genes in water-stress and irrigated conditions were calculated with the help of available RNA-seq data. Differential gene expression was calculated for each of the candidate genes under drought and control. Fifty DEGs in Banglami and seventy-two DEGs in Ranjit having a Log2 Fold change greater than ± 2.5 up to ± infinity were considered significant. Few of the differentially expressed genes having a significant Log2Fold change as revealed by the sequencing data, were confirmed experimentally under drought situation using qRT-PCR. Stress-responsive candidate genes involved in nitrate uptake, DNA repair, ubiquitination, enzymatic activities, cell cycle activities, etc. which can play a significant role in conferring drought tolerance were identified in the drought-related QTLs. The expression pattern of the candidate genes in qRT-PCR was found similar to transcriptome studies, signifying important role of the candidate genes in drought tolerance mechanism in rice.

Theses

Browse

Thesis Type

Type

Author

Date

Subject

Search Results