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Dr. Rajendra Prasad Central Agricultural University, Pusa

In the imperial Gazetteer of India 1878, Pusa was recorded as a government estate of about 1350 acres in Darbhanba. It was acquired by East India Company for running a stud farm to supply better breed of horses mainly for the army. Frequent incidence of glanders disease (swelling of glands), mostly affecting the valuable imported bloodstock made the civil veterinary department to shift the entire stock out of Pusa. A British tobacco concern Beg Sutherland & co. got the estate on lease but it also left in 1897 abandoning the government estate of Pusa. Lord Mayo, The Viceroy and Governor General, had been repeatedly trying to get through his proposal for setting up a directorate general of Agriculture that would take care of the soil and its productivity, formulate newer techniques of cultivation, improve the quality of seeds and livestock and also arrange for imparting agricultural education. The government of India had invited a British expert. Dr. J. A. Voelcker who had submitted as report on the development of Indian agriculture. As a follow-up action, three experts in different fields were appointed for the first time during 1885 to 1895 namely, agricultural chemist (Dr. J. W. Leafer), cryptogamic botanist (Dr. R. A. Butler) and entomologist (Dr. H. Maxwell Lefroy) with headquarters at Dehradun (U.P.) in the forest Research Institute complex. Surprisingly, until now Pusa, which was destined to become the centre of agricultural revolution in the country, was lying as before an abandoned government estate. In 1898. Lord Curzon took over as the viceroy. A widely traveled person and an administrator, he salvaged out the earlier proposal and got London’s approval for the appointment of the inspector General of Agriculture to which the first incumbent Mr. J. Mollison (Dy. Director of Agriculture, Bombay) joined in 1901 with headquarters at Nagpur The then government of Bengal had mooted in 1902 a proposal to the centre for setting up a model cattle farm for improving the dilapidated condition of the livestock at Pusa estate where plenty of land, water and feed would be available, and with Mr. Mollison’s support this was accepted in principle. Around Pusa, there were many British planters and also an indigo research centre Dalsing Sarai (near Pusa). Mr. Mollison’s visits to this mini British kingdom and his strong recommendations. In favour of Pusa as the most ideal place for the Bengal government project obviously caught the attention for the viceroy.

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2016 - 201811
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Subject: Agricultural Biotechnology × End date: 2018 × Start date: 2016 × Type: Thesis × Thesis Type: M.Sc ×
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Now showing 1 - 9 of 11
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    ThesisItemOpen Access
    Candidate gene markers based molecular profiling for grain zinc accumulation in rice
    (Dr. Rajendra Prasad Central Agricultural University, Pusa, Samastipur, 2018) Kumari, Kumkum; Sharma, V.K.
    A study was conducted to determine the genetic variation and divergence in relation to grain zinc accumulation amongst rice varieties using candidate gene based panel of reported primers and to examine the genetic importance of zinc transporter candidate gene based panel of designed primers in discrimination for differential zinc accumulation amongst rice varieties. Twenty-eight locally adapted varieties and advanced breeding lines of rice were evaluated in randomized block design with three replications and the seeds collected after harvesting of the crop were utilized for determination of zinc content in unpolished grains. Eighteen entries selected from the two extremes of grain zinc distribution range constituted the final experimental material and utilized during molecular characterization. Genomic DNA was extracted from two to three weeks old seedlings of purposefully selected set of 18 varieties and then targeted amplification of the genomic DNA was achieved by using a panel of 14 candidate gene specific 14 reported primers and 14 designed primers. Exploitable extent of variability was observed with respect to grain zinc accumulation amongst the set of 28 rice varieties initially evaluated as experimental materials. Zinc content, which varied from 8.18 ppm to 21.53 ppm, was found to be considerably higher in unpolished grains of RAU 3036, Sanwal Basmati, Rajendra Nilam and Rajendra Bha gwati. Using a panel of 14 candidate genes specific 14 reported primer pairs, reproducible amplification was successfully achieved with 12 primer pairs amongst which only eight primer pairs generated polymorphic amplified products. Successful amplification with two candidate genes specific reported primers, namely, OsNAC and OsNRAMP6a was notachieved. Contrarily, each of the 14 designed primer pairs exhibited reproducible amplification, but polymorphic amplified products were generated with only eight primer pairs. Appearance of amplified products in the form of bands at different positions on the gel revealed differential migration due to differences in overall size of the products generated from targeted amplification of specific region of genome. Molecular level genetic polymorphism among the entries was recognized on the basis of variation in respect of position of bands. Ample genetic differentiation and divergence was revealed at the molecular level amongst the rice varieties subjected to molecular characterization using the candidate genes specific and polymorphic panels of reported as well as designed primer pairs. Results from reported primers and designed primers based analysis were in well agreement with each other. Furthermore, hierarchical classification pattern of rice varieties was almost completely corroborated by principal coordinate analysis based spatial distribution pattern of genetic profiles of rice varieties. Hierarchical cluster analysis as well as principal coordinate analysis based on a combination of polymorphic and informative eight reported and eight designed primer pairs provided better expression of differentiation and divergence amongst the rice varieties subjected to molecular characterization. Thus, the use of 14 candidate genes specific 16 polymorphic markers in the genetic analysis exhibited a remarkably higher level of genetic polymorphism, which allowed unique genotyping of eighteen entries included in the analysis. Hence, these markers can be effectively and efficiently utilized for grain zinc accumulation related discrimination of rice genotypes and selection of parental genotypes for genetic improvement in relation to grain zinc biofortification. Microsatellites were detected within the candidate genes and within the amplicons, thereby providing a basis to deduce that the variation present in candidate genes, as observed in terms of differences in the molecular size of the genomic regions spanned by the primer pairs, may be a role player in the differential grain zinc accumulation in rice varieties. Single marker analysis established the association of four markers, namely, OsNACK, OsZIP1-1, OsNRAMP7 and OsNRAMP7K with grain zinc accumulation. These four markers can be effectively used in marker-assisted selection program for grain zinc biofortification in rice. Inter-crossing diverse genotypes from different clusters can lead to successful pyramiding of desirable alleles through molecular breeding program. Parental genetic diversity will undoubtedly increase the probability of identifying desirable recombinants during screening for improvement in relation to grain zinc biofortification.
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    ThesisItemOpen Access
    Genome scanning for differential grain iron accumulation in rice using candidate gene markers
    (DRPCAU, Pusa, Samastipur, 2018) Arjun, Kale Rohan; Sharma, V.K.
    Analysis of functional polymorphic markers provide ace in diversifying the selective genotypes that may stand useful in biofortification programme for improving the genotypes in relation to their micronutrient composition. A study was conducted based on this consideration to evaluate the molecular level genetic divergence and to investigate the genomic regions and genes associated with natural variation of grain iron concentration among purposefully selected landraces, advanced derivatives and improved varieties of rice. With this objective, 28 locally adapted landraces, improved varieties and advanced breeding lines were evaluated in randomized block design with three replications and the grains collected after harvesting of the crop were utilized for determination of iron content in unpolished grains. The digestion was carried out by di-acid mixture which included nitric acid and perchloric acid (9:4) and iron content was determined with the help of atomic absorption spectrophotometer. Eighteen entries selected from the two extremes of grain iron distribution range were utilized during molecular characterization. Genomic template was extracted from two to three weeks old seedlings of these purposefully selected varieties and then targeted amplification of the genomic regions was achieved by employing a panel of six candidate genes specific 18 primers and a panel of candidate genes based 12 microsatellite primers. Candidate gene specific primers were designed using primer blast tool for finding specific primers sequence of the identified candidate gene and appropriate primer sequences were selected for their utilization during molecular profiling. Exploitable genetic variability was observed in relation to grain iron concentration amongst 28 locally adapted landraces, varieties and advanced breeding lines of rice. Using the standard deviation for the range of variation (16.39 to 39.04 ppm) as the criterion, the rice varieties were classified into very low, low, moderate, high and very high grain iron containing groups. Putative candidate genes under investigation in the present study were searched for the presence of microsatellite sequences within the candidate gene sequences using microsatellite identification tools. Altogether 40 microsatellites were detected within the six putative iron transporter candidate genes. These microsatellites had dinucleotide to hexanucleotide repeat motifs. Only two microsatellites were detected in OsNRAMP1 and OsZIP8, whereas 18 microsatellites were detected in OsNRAMP5. Experimental results provided a basis to deduce that the variation present in candidate genes, as revealed in terms of differences in the molecular size of the genomic regions spanned by the primer pairs, may be a role player in the differential grain iron accumulation in rice varieties. Using a panel of six candidate genes specific 18 primers and 12 microsatellite primers, reproducible amplification was successfully achieved in the purposefully selected rice varieties. All the primers generated polymorphic amplified products. While each one of the eight polymorphic candidate gene specific primers, namely, APRT1a, APRT1b, APRT1c, OsNAC5a, OsNAC5c, OsNRAMP1c, OsZIP10a and OsZIP10c, generated three allelic variants, each one of the remaining ten polymorphic primers, namely, OsNAC5b, OsNRAMP1a, OsNRAMP1b, OsNRAMP5a, OsNRAMP5b, OsNRAMP5c, OsZIP8a, OsZIP8b, OsZIP8c and OsZIP10b, detected two allelic variants. Differential amplification pattern was also exhibited by candidate genes specific 12 microsatellite primers. While some of the primers generated several markers, some generated only few allelic variants. Altogether 72 allelic variants were detected among the 18 entries with an average of 6.0 alleles per primer. Polymorphic information content of candidate gene specific primers ranged from 0.278 to 0.710 with an average value of 0.483, while that of candidate genes specific microsatellite primers ranged from 0.154 to 0.864 with an average of 0.690. Sizable molecular level genetic differentiation and divergence was revealed amongst the rice varieties using the candidate gene specific primers as well as candidate genes specific microsatellite primers. Hierarchical classification pattern based on similarity coefficient matrix of pair-wise combinations of entries, which were accommodated into different clusters, was highly consistent with principal coordinate analysis based spatial distribution pattern of genetic profiles. Hierarchical cluster analysis as well as principal coordinate analysis using candidate gene markers as well as candidate genes specific microsatellite markers enabled differentiation and classification of entries with remarkably higher level of consistency in relation to their grain iron concentration. Hence, these markers can be effectively and efficiently utilized for discrimination of rice genotypes and selection of parental genotypes for genetic improvement in relation to grain iron biofortification. Single marker analysis established the association of two candidate genes specific markers (APRT1c and OsNRAMP1b) and candidate gene specific four microsatellite markers (OsNAC5A, OsNAC5B, OsNRAMP1A and OsZIP10A), with grain iron concentration. These six markers can be effectively utilized in selection program for grain iron biofortification in rice.
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    ThesisItemOpen Access
    Molecular characterization of SS-III gene promoter with respect to Heat tolerance in Wheat
    (Dr. Rajendra Prasad Central Agricultural University, Pusa, Samastipur, 2018) Kumar, Devendra; Kumar, Rajeev
    Bread wheat (Triticum aestivum L.) is one of the most important staple food grains of the human race as it fulfils the significant proportion of dietary calories requirements of the world population. Starch is a major component of wheat flour, accounting for 65-70% of the dry weight of the wheat grain. There are 6 distinct classes of starch synthase: GBSS, SSI, SSII, SSIII, SSIV and SSV involved in the synthesis of starch. SS genes containing promoter of 2kb sequence were retrieved in wheat and other crop species using by the bioinformatics tools. The promoter sequence of the reference crop species or wheat was used as a query in the cereals database. 68 cis-acting regulatory elements and 118 total number of motif sequences were found across the species under study. A maximum number of motifs (51) were found in Amaranths spp. In wheat total, 27 motifs were found. ERE (Ethylene Responsive Element) was found in SS-III in wheat. G-box (cis-acting regulatory element involved in light responsiveness) and MBS (MYB binding site involved in drought-inducibility) were found in all the species of monocots and dicots. Four novel conserved motifs e.g. ARE (anaerobic induction); GC-motif (anoxic specific inducibility); TGA (auxin-responsive element); TGACG motif (MeJA responsiveness) were found in the SSIII gene promoter consensus sequence obtained through wet lab experiment but found absent in the retrieved sequences. In Phylogenetic analysis Maximum genetic similarities were observed for Phaseolus SS-I & Phaseolus SS-II and Arabidopsis SS-II & Arabidopsis SS-I respectively where-as maximum dissimilarities was observed between Phaseolus vulgaris L. SSII and Brachypodium SS-II. Wheat SS-I was found more closely related to its SS-III, SS-II was found closely related to Brachypodium SS-III, and SS-IV promoter was found closely related to SS-III of Zea mays. We identified a total of 72 SNPs were identified over a total region of 1496 bp. Out of 72, 24 SNPs are of transition types and 48 SNPs are of transversion types in the studied genotypes of wheat with different degree of sensitivity towards heat stress. In heat sensitive genotypes (PBW-343, SISD, HD-2967, and Sonara-64) there were 29, 12, 6, and 1 specific SNPs identified respectively. 7 and 17 specific SNPs were also identified in heat tolerant genotypes (KSG-1186) and (Ipecarabe) respectively. In heat tolerance genotypes (KSG-1186) and (Ipecarabe) identified more transversion types (A/T) SNP than transition types. Analysis of types of substitution among the identified SNPs revealed far more abundance of transversion than transition and the ratio of transition to transversion (Ts/Tv) as 0.5.
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    ThesisItemOpen Access
    Molecular evaluation of sterility maintainers and fertility restorers for wild abortive rice cytoplasm
    (Dr. Rajendra Prasad Central Agricultural University, Pusa, Samastipur, 2018) Kumar, Pradeep; Sharma, V. K.
    A set of forty-five test crosses involving two wild abortive type cyto-sterile lines and twenty-eight pollen parents was evaluated to assess pollen and spikelet fertility restoration for identification of sterility maintainers and fertility restores of wild abortive cyto-sterility system in rice. Using a panel of thirty-two microsatellite primers, genetic polymorphism at molecular level was examined for characterization and differentiation of sterility maintainers and fertility restorers identified among pollen parents. Analysis of pollen and spikelet fertility in test crosses clearly reflected the sterility maintaining ability of pollen parents TCP-134-2, TCP-150-1, TCP-188-3 and TCP-185-3. Similarly, the pollen parents TCP-17-1, TCP-28-3, TCP-86-1, TCP-174-4, TCP-104-1, TCP-183-1, TCP-190-3, TCP-191-3 and TCP-193-3 were identified as effective fertility restorers. Altogether 256 allelic variants including 112 unique alleles and 144 shared alleles were identified with an average of 8.0 alleles per primer using a panel of 32 primer pairs covering all the chromosomes. The primer targeting tri-nucleotide and di-nucleotide repeat motifs, in general, detected more allelic variants than primers targeting tetra-nucleotide repeat motifs. Further, primers with GA, TG, AC, CT and TA di-nucleotide repeat motifs detected more allelic variants than the primers with TC and AG di-nucleotide repeat motifs. The primer pairs RM 216, RM 6100, RM 280, RM 3873, RM 10313, RM 558, RM 250, RM 283, RM 171, RM 3233, RM 341, RM 427, RM 206, RM 5373, RM 152, RM 591, RM 524 and RM 17 appeared to be highly polymorphic and comparatively more informative for the purpose of molecular profiling of entries. Analysis of divergence pattern allowed discrimination of effective fertility restorers from partial fertility restorers and complete sterility maintainers. Clustering pattern of the entries based on hierarchical classification, neighbour joining tree and principal co-ordinate analyses yielded more or less similar results. Allelic diversity data generated from amplification pattern of the six fertility restoration related primer pairs, namely, RM 591, RM 1108, RM 3233, RM 3873, RM 6100 and RM 8146, also unambiguously discriminated eight effective fertility restorers from two partial fertility restorers and four complete sterility maintainers. Therefore, these six primer pairs were validated with sufficiently greater efficiency (94.4%) for identification of sterility maintainers and fertility restorers of wild abortive type cyto-sterility sterility system in rice.
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    ThesisItemOpen Access
    Molecular Characterization of Aromatic Rice Genotypes using Microsatellite Markers
    (Dr. Rajendra Prasad Central Agricultural University, Pusa, Samastipur, 2018) Priyadarshini, Manisha; Sharma, V. K.
    A study was conducted to examine the nature and extent of genetic differentiation and divergence among the aromatic rice genotypes using microsatellite markers.The materials were grown in petriplates for extraction of genomic DNA from the young seedlings and then targeted amplification of the genomic DNA was achieved by using a panel of eighteen microsatellite based primer pairs covering four chromosomes in the genome of rice. The statistical methods and parameters used for deriving inference were polymorphism per cent, polymorphism information content, discrimination coefficient, similarity coefficient, principal coordinate analysis and numerical taxonomic analysis of divergence. Amplification was successfully achieved with all the microsatellite primers used in the present study. Appearance of bands at different positions on the gel revealed differential migration of amplified products due to differences in overall size of the products generated from targeted amplification of specific region of genome. The polymorphism among the entries was recognized on the basis of presence or absence of bands, in addition to variation in respect of number and position of bands.Altogether 123 allelic variants were detected among the eighteen aromatic rice entries with an average of 4.92 alleles per locus.The number of allelic variants per primer pair ranged from three in the case of E03.92.0 to ten in the case of RM 252, RM 256, RM 284 and RM 7356. A total of 76 shared and 47 unique allelic variants were generated amongst the entries.The primer pairs RM 7356, RM 252, RM 284, RM 223, RM 444, Aro7, E11.44.5, RM256 and RM 505 generated considerably greater percentage of unique alleles. Appearance of more than one band in the same entry indicated the existence of the duplicated region in the genome. The primer pairs RM 223, RM 252, RM 444, RM 505, RM 252, RM 7049 and RM 7356 generated amplified products due to amplification of more than one locus. Considering the magnitude of polymorphic information content value in conjunction with number of alleles and polymorphism per cent, the primer pairs RM 256, RM 284, Aro7, RM 223, RM 252, RM 444, RM 7356appeared to be highly informative.Occurrence of null allele for a microsatellite locus associated with primer pairs RM 505 was noticed reflecting null allele in combination with one of the eighteen entries under evaluation.Relatively higher magnitude of discrimination coefficient was obtained for the primer pairs RM 256, RM 284, Aro7, RM 8264, RM 223, RM 252, RM 444, RM 7356, RM 7049, ARSSR-3 and RM 515, indicating their greater efficiency in respect of their ability to unambiguously discriminate the pair-wise combination of aromatic rice entries. Microsatellite loci with tetra-nucleotide repeat motifs tended to detect greater number of alleles than the repeat loci withdi-nucleotide, tri-nucleotide and complex repeat motifs.Any relationship between the repeat number and the number of identified alleles was not observed in the present study.Simple sequence repeat loci with CT, GA and AT di-nucleotide repeat motifs tended to detect comparatively greater number of allelic variants. Ample genetic variation at molecular level amongst the aromatic rice entries under evaluation was inferred on the basis of the estimates of similarity coefficients. Considering the broad classification of entries, the entries appeared to bebasically divided into three groups, which were further divided into five clusters to allow the entries with relatively more similar pattern for markers to be clustered together. Principal coordinate analysis based two dimensional plots of the genetic profiles supported the results of hierarchical classification based grouping of entries.. Microsatellite marker based analysis revealed unique or entry specific allele which could be useful as molecular fingerprints in the unambiguous discrimination and identification of aromatic rice genotypes. The use of eighteen microsatellite markers in the analysis of aromatic rice entries exhibited a remarkably higher level of genetic polymorphism, which allowed unique genotyping of eighteen entries included in the analysis.
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    ThesisItemOpen Access
    Characterization of gene(s) involved in starch synthesis with respect to heat stress tolerance in wheat
    (Dr. Rajendra Prasad Central Agricultural University, Pusa (Samastipur), 2018) Mishra, Bhavya Priyadarshini; Kumar, Rajeev
    High temperature stress during the reproductive and grain filling phase is considered detrimental for wheat yield. In the present study thirty-two wheat (Triticum aestivum L.) genotypes were characterized with respect to heat stress under three different sowing conditions viz. Timely sown (S1), Late sown (S2) and very Late sown (S3) along with detailed structural, functional and molecular characterization of Starch Synthase III gene which is most critical for grain development in Wheat. The experimental materials were evaluated in randomized block design (RBD). The lowest performance of the genotypes were recorded under very late sowing condition (S3) . The impact of heat stress with respect to performance of the genotypes was found significantly higher in case of very late sown condition than in late sown stress condition. Genotypes C-306, Chiriya-3, Dharwad Dry,IC532653, Giza 155, Raj-4083, HI-1563, Cus/9/Prulla and DBW 14 exhibited stable performance for most of the traits whereas PBN -51, PBW 343, Sonora 64, Tepeko, Seri-82, BWL-1793 showed least stable performance in in all the three growing conditions.Starch Synthase (SS) has five distinct isoforms of which SSIII is involved in the formation of longer glucan chain length and most critical for grain filling. Here we also report identification and detailed characterization of ‘true’ orthologs of the well-characterized maize SSIII (ZmSSIII), among six monocots and two dicot species. ZmSSIII orthologs have nucleotide sequence similarity ranging from 56-81%. Variation in gene size among various orthologs ranged from 5.49 kb in Arabidopsis to 11.62 kb in Brachypodium and the variation was mainly due to intron size and indels present in the exons 1 and 3. Number of exons and introns were highly conserved among all orthologs however. While the intron number was conserved, intron phase showed variation at group, genera and species level except for intron 1 and 5. Several species, genera, and class specific cis-acting regulatory elements were identified in the promoter region. Despite of significant sequence variation among orthologs, most of the motifs and their relative distances are highly conserved among the orthologs. Homeologs of wheat SSIII gene showed tissue and developmental stage specific expression pattern with the highest expression recorded in developing grains in eight genotypes under consideration. Three different homeo-loci, for SSIII on chromosome group 1 showed dramatically different expression pattern during different grain developmental stages and exhibited low expression in stress condition in most of the genotypes except for KSG151, KSG053 and KSG270 that had stable expression under stress conditions.
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    ThesisItemOpen Access
    Gene Expression Analysis For Biosynthesis And Catabolism Of Glucosinolates In Rocket (Diplotaxis tenufolia L.) Under Abiotic Stress
    (Dr. Rajendra Prasad Central Agricultural University, Pusa (Samastipur), 2017) Mishra, Shubhi; Kumar, M.
    Glucosinolates are one of the important nutritive content of rocket (Diplotaxis tenufolia L.), a leafy vegetable salad crop. They are responsible for its pepper taste, pungent aroma and medicinal properties. In this experiment, content of chlorophyll and chlorophyll fluorescence parameters were measured and biochemical analysis of glucose and glucosinolates content was done. The experiment was further followed by expression profiling of the key genes of metabolic pathway of glucosinolate using real-time PCR. The results indicated that the plant had some adaptation mechanism to mitigate the adverse effects of heat and salt stress which resist the changes in the chlorophyll content and chlorophyll fluorescence of rocket leaves as non-significant changes in the chlorophyll content and chlorophyll a fluorescence of rocket leaves.An increase in glucosinolate content was observed due to heat and salt stress in agreement with the inference derived on the basis of gene expression analysis. However in the case of roots, it appeared that glucosinolates content started decreasing after reaching the saturation point which was amply supported by higher level of expression of glucosinolates catabolism gene DtMyrosinase as observed in roots.It appeared that glucosinolates content can be increased if grown under heat and salt stress and can also be limited if grown under controlled condition. Isolation of the full length of key genes involved in glucosinolates biosynthesis namely, DtCytochromeP79F1 and catabolism namely, (DtThiol-methyl transferase) was also done through RACE. Altogether ten primers including eight primers specific to target the eight genes of glucosinolate metabolic pathway and two RACE specific primers were designed and validated during the present study which may be further utilized as tools to examine target region amplification polymorphism and expression in relation to glucosinolate biosynthesis pathway in rocket plants.
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    ThesisItemOpen Access
    Genome wide association study for identification of genes related to ion accumulation in rice grain
    (Dr. Rajendra Prasad Central Agricultural University, Pusa (Samastipur), 2017) Kumar, Ajay; Sharma, V. K.
    A study was conducted to investigate and identify the candidate genes in relation to cadmium and zinc transportation and accumulation in rice. Using 281 diverse rice accessions and employing genome wide association analysis to relate 46294 single nucleotide polymorphic panel and cadmium and zinc ionomic data, mixed linear model was adopted to establish the association, if any, between genotypic and ionomic data. Statistical tools used and computational applications adopted during the analyses include Discriminant Analysis of Principal Components (DAPC), Principal Coordinate Analysis (PCoA), Neighbour Joining Tree, Structure, Haploview and R-tool. Differential accumulation of cadmium and zinc in different vegetative tissues of two diverse rice accessions was also investigated under cadmium induced stress condition. Finally, expression profiles of OsHMA2, OsHMA3 and OsPCSs genes under cadmium induced stress condition were examined. Differential cadmium accumulation was recorded in grains of rice accessions evaluated under alternate wetting and drying and permanent flooding conditions as well as across the conditions. While in the case of zinc, differential accumulation was observed between accessions but differences recorded between conditions were not so pronounced. PCoA based on SNPs data derived using 297 accessions clearly indicated that 12 entries were very diverse from rest of the entries under evaluation. Further, an analysis using neighbour joining tree based on distance matrix also revealed that these 12 genotypes were remarkably distant from rest of the entries. Cluster analysis applied to detect population genetic structure clearly indicated that 10 out of these 12 accessions belonged to a separate cluster. Finally, an analysis carried out after exclusion of 16 accessions including 12 accessions, which were found to be more diverse based on PCoA and neighbour joining tree and 4 very late maturing accessions, established that remaining 281 accessions belonged to three clusters. DAPC employed to find out the optimum number of clusters confirmed the results obtained through PCoA, neighbour joining tree and cluster analysis applied to detect population genetic structure in the present investigation. Association analysis revealed twelve and two markers; seven and one candidate loci; and 109 and 10 candidate genes associated with cadmium and zinc accumulation, respectively. Haplotype analysis of polymorphic markers identified in relation to cadmium and zinc accumulation, therefore, resulted in allocation of seven and one loci, respectively, in question to haplotype groups. Result of the present study led to the identification of candidate genes involved in cadmium and zinc accumulation under aerobic condition. Altogether, eight and five genes appeared to be involved in cadmium and zinc accumulation, respectively. Present study, therefore, provided the evidence based on which the candidate genes most likely to be involved in zinc accumulation have been proposed. Most probable involvement of MATE and PEZ family genes in cadmium accumulation in rice grain was confirmed on the basis of results of present study. Cadmium accumulation property appeared less dependent on its uptake by the plants but more dependent on the capacity of the accession to trap the cadmium into the roots by synthesizing two folds non-protein thiol content, thereby preventing its translocation or mobility from root to shoot. Antagonistic effect of cadmium accumulation was observed in respect of zinc accumulation in rice accessions. Expression analysis in relation to cadmium ion translocation reflected that OsHMA3 was more expressed in less efficient accession as compared to the efficient accession. Therefore, the overexpression of OsHMA3 most probably resulted in low accumulation of cadmium in the plants of less efficient accession due to lesser translocation.
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    ThesisItemOpen Access
    Microsatellite markers based characterization of rice genotypes in relation to drought tolerance
    (Dr. Rajendra Prasad Central Agricultural University, Pusa (Samastipur), 2017) Choudhary, Shuchi Shankar; Kumar, Mithilesh
    Eighteen genotypes of rice were evaluated for various physio-morphological characters and to investigate the microsatellite markers based polymorphism for their characterization and differentiation using polymorphic and informative markers in order to estimate the extent of genetic diversity among these rice genotypes.Experimental materials were evaluated in randomized block design with three replications during three consecutive years. Molecular characterization was done by targeted amplification of the genomic DNA using a panel of thirty two microsatellite primer pairs covering nine chromosomes. Statistical methods and parameters used for deriving inference were analysis of variance, range, mean values, relative mean performance, drought tolerance indices, variability parameters, correlation andpath coefficients. An analysis of variance revealed significant differences among the genotypes for all the characters evaluated during present study. Considerable extent of variability existed for all the attributes recorded among the genotypes. The coefficient of variability was recorded moderate to high genetic variability for number of grains per panicle, root length and yield per plant under normal condition consistently over years, whereas for leaf rolling, number of tillers per plant, number of grains per panicle, plant height, harvest index and yield per plant under drought condition. Yield per plant recorded high variability amongst the genotypes evaluated under both conditions.The genotypes RAU-1426-4-3-2-57-2, RAU-1416-4-2-5-2-2 and Sahbhagi Dhan recorded remarkably higher mean performance fornumber of tillers per plant, number of grains per panicle, 1000-grains weight, harvest index and yield per plant, under both drought and normal conditions. Similarly, genotypes like RAU-1397-2-5-8-1-2-5-4, RAU-1453-12, RAU-1451-6-6-1-1-5-2 and Sahbhagi Dhan exhibited superior performance in respect of root length, plant height, dry root weight, relative leaf water content, canopy temperature and number of grains per panicle under both drought and normal conditions. The relative mean performance of genotypes, namely,RAU-1421-12-1-7-4-3, RAU-1415-3-5-7-6-9-5-3, RAU-1401-1-8-1-5, RAU-1478-5-2-2-4-6, RAU-1451-6-6-1-1-5-2, RAU-1416-4-2-5-2-2, RAU-1426-4-3-2-5-7-2, RAU-1453-12 and Sahbhagi Dhan to be relatively more drought tolerant than other genotypes under evaluation. Root length, plant height, dry root weight, spikelet fertility, number of grains per panicle, 1000-grain weight, harvest index and yield per plant exhibited moderate to high heritability in broad sense coupled with moderate to high estimates for genetic advance as per cent of mean under both drought and normal conditions over years.Correlation studies, revealed that root length, number of tillers per plant, relative leaf water content, spikelet fertility, 1000-grain weight and harvest indexshowed significant positive association with yield per plant under both conditions. However, positively significant association was also observed between yield per plant and number of grains per panicle under drought condition.Path coefficient analysis revealed that the characters, namely, numbers of tillers per plant, root length, dry root weight1000-grain weight and harvest index had considerable positive direct effects on yield per plant more or less consistently across the years under drought as well as normal conditions.Four tolerance indices viz. TOL, SSI, DTE and STI were used. According to TOL, SSI and DTE, 12 genotypes,namely, RAU-1421-12-1-7-4-3, RAU-1415-3-5-7-6-9-5-3, RAU-1401-1-8-1-5, RAU-1478-5-2-2-4-6, RAU-1477-9-7-2-2-5-7-3, RAU-1451-6-6-1-1-5-2, RAU-1416-4-2-5-2-2, RAU-1426-4-3-2-5-7-2, RAU-1463-16, RAU-1471-10, RAU-1453-12 and Sahbhagi Dhan appeared to be relatively more tolerant to drought stress. Among these 12 genotypes, 7 genotypes namely,RAU-1421-12-1-7-4-3, RAU-1415-3-5-76-9-5-3, RAU-1451-6-6-1-1-5-7-3, RAU-1416-4-2-5-2-2, RAU-1426-4-3-2-5-7-2, RAU-1453-12 and Sahbhagi Dhan recorded higher STI valueexhibiting high tolerance to drought condition. Using a set of thirty two microsatellite primers generated allelic variants ranged from 4 in case of RM 85 to 16 in case of RM 324 and RM518.The primer pair RM 3,RM 18, RM 72, RM 87, RM225, RM 262, RM302, RM 324, RM 327, RM 416, RM 518, RM 520, RM 521, RM1349, RM 5443 and RM 23679generated amplified products due to amplification of more than one microsatellite locus. Altogether 300 allelic variants were detected at 48 microsatellite loci with an average of 6.25 alleles per locus. A total of 174 shared and 126 unique allelic variants were generated. Considering the number of alleles in conjunction with the level of polymorphism detected, the primersRM 231, RM 225, RM 163, RM 555, RM 212, RM 72, RM 3549, RM 1349, RM 521, RM 5443 and RM 302 appeared to be more informative primers.Analysis of divergence pattern based on microsatellite markers allowed differentiation and classification of rice genotypes into two groups. The first multi-genotypic group consisted of twelve genotypes, namely, RAU-1428-6-7-3-6, RAU-1417-2-1-5-7-7,RAU-1421-12-1-7-4-3, RAU-1415-3-5-76-9-5-3, RAU-1401-1-8-1-5, RAU-1428-4-3-2-7-26, RAU-1478-5-2-2-4-6, RAU-1477-9-7-2-2-5-7-3, RAU-1451-6-6-1-1-5-2, RAU-1416-4-2-5-2-2, RAU-1397-2-5-8-1-2-5-4 and RAU-1451-3-5-7-6-9-5-1 whereas the second multi-genotypic group consisted of six genotypes, namely, RAU-1426-4-3-2-5-7-2, RAU-1428-3-1-5-4, RAU-1463-16, RAU-1471-10, RAU-1453-12 and Sahbhagi Dhan.The maximum similarity value of 0.46 was observed between the genotypes i.e. genotype RAU-1477-9-7-22-5-7-3 and RAU-1451-6-6-1-1-5-2 indicating that these were more closely related. The panel of microsatellite markers employed for molecular characterization in the present study showed very high degree of efficiency in discrimination of genotypes in relation to drought tolerance.