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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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ThesisItem Open Access Molecular characterization of rice genotypes using salt stress responsive candidate gene based markers(DRPCAU, Pusa, 2020) Agarwal, Ruchi; Sharma, V.K.A study was undertaken for characterization of salt tolerance related response of 18 rice genotypes using morpho-agronomic characters and salt stress responsive 12 candidate gene based newly designed markers. Evaluation of genotypic response to salt stress at seed germination and early seedling stage was performed at 4 dSm-1 and 8 dSm-1 EC levels along with control by adopting a completely randomized design with two replications. Pot experiment was carried out at 8 dSm-1 and 12 dSm-1 EC levels given at vegetative and reproductive stages, respectively, along with control in completely randomized design with two replications. Field evaluation was conducted in randomized block design with two replications under normal and salt affected field conditions. The genotypes were further characterized at molecular level using 12 candidate gene based newly designed 23 genic primers and 21 genic microsatellite primers. Differential genotypic response to salt stress was noticed at germination stage and early seedling stage. Based on response exhibited at early seedling stage, the genotypes CSR-13, CSR- 23, CSR-27, CSR-30, CSR-36, CST7-1 and CSR-2K-262 were characterized with considerably higher level of tolerance to salt stress in comparison to the genotypes NDRK-11-1, NDRK-11-3, NDRK-11-4, NDRK-11-5, NDRK-11-6, NDRK-11-7, CSR-2K- 219 and CSR-2K-242, which were found to be moderately tolerant. The remaining three genotypes, namely, IR-36, IR-64 and Swarna, were found to be susceptible to salt stress. Principal component analysis based spatial distribution pattern of the genotypes in two dimensional projections distinctly separated the genotypes into two broad groups. The first group was further divided into two sub groups consisting of six salt tolerant genotypes and nine closely located moderately tolerant genotypes, whereas the second multi-genotypic group consisted of three salt susceptible genotypes. The basic pattern of differentiation and interrelationships among the genotypes was found to be similar in dendrogram. A comparison of the relative mean values of the genotypes evaluated for 14 morpho-physiological attributes, such as, panicle length, panicles per plant, spikelets per panicle, filled grains per panicle and unfilled grains per panicle, 100-seed weight, biological yield, grain yield per plant, root length, root volume, root dry weight, relative water content, SPAD value and K/Na ratio in pot experiment was made with mean index (MI) value. Based on the results, the genotypes CSR-13, CSR-23, CSR-27, CSR-30, CSR-36 and CST7-1 were found to be salt tolerant, whereas NDRK-11-1, NDRK-11-3, NDRK-11-4, NDRK-11- 5, NDRK-11-6, NDRK-11-7, CSR-2K-219, CSR-2K-242 and CSR-2K-262 were rated as moderately tolerant and genotypes IR-36, IR-64 and Swarna were found susceptible. Spatial distribution pattern of the genotypes in two dimensional ordinations based on principal component analysis broadly divided the genotypes into two groups, efficiently separating tolerant and moderately tolerant genotypes from susceptible genotypes. The pattern of genotypic discrimination was similar corroborated by dendrogram. Tolerance indices, such as, TOL, MP, GMP, SSI and STI based principal component analysis and hierarchical cluster analysis also discriminated the susceptible genotypes from moderately tolerant and tolerant genotypes. Relative mean performance of genotypes evaluated under normal and salt affected field conditions for 11 morpho-agronomic characters also reflected that the genotypes CSR- 13, CSR-23, CSR-27, CSR-30, CSR-36, CST7-1 and CSR-2K-262 exhibited tolerant response, whereas NDRK-11-1, NDRK-11-3, NDRK-11-4, NDRK-11-5, NDRK-11-6, NDRK-11-7, CSR-2K-219 and CSR-2K-242 were found moderately tolerant and the genotypes IR-36, IR-64 and Swarna were rated as susceptible. Principal component analysis based spatial distribution pattern separated the genotypes into two broad groups, separating the three susceptible genotypes from moderately tolerant and tolerant genotypes. A similar classification pattern was revealed by average taxonomic distance based dendrogram. Recognizable molecular level genetic polymorphism amongst the 18 genotypes was revealed by amplification of genomic template using 12 candidate gene specific 23 newly designed primer pairs. All primer pairs generated 135 allelic variants with an average of 5.86 alleles per primer. The PIC values exhibited significant variation with the values ranging from 0.308 in primer pair DREB1F-1 to 0.905 in primer pairs SHMT1-1. Considering the level of polymorphism detected, the candidate gene based primer pairs OsHKT2;3-1, OsHKT2; 3-2, SNAC1-1, SNAC1-2, CDMK-1, CDMK-2, DUF6-2 SHMT1- 1, SHMT1-2 and SHMT2-1 appeared to be highly polymorphic and more informative primers for the purpose of discrimination of rice genotypes in relation to salinity tolerance. Principal coordinate analysis and hierarchical classification unambiguously discriminated the tolerant and susceptible genotypes, reflecting the usefulness of these newly designed markers. Using major ion transporter, transcription factor and oxidative metabolic pathway related candidate gene specific newly designed 21 microsatellite primer pairs, ample molecular level genetic polymorphism was detected amongst 18 genotypes. All primer pairs generated 131 allelic variants with 6.23 alleles per primer. The number of unique and shared alleles varied from 0 to 5 and 1-6, respectively. The PIC values ranged from 0.706 to 0.955. Taking into consideration the measures of polymorphism, the primers OsHKT1;5-B, OsHKT1;5-D, OsHKT2; 3-A, OsHKT2; 4–A, DREB1F-C, CDMK-B, DUF6-B, SHMT1- B, and SHMT2-A were found as relatively more polymorphic. Spatial distribution pattern of genotypes based on principal coordinate analysis of the genetic profiles generated by these newly designed primers as well as hierarchical classification unambiguously differentiated the tolerant and susceptible genotypes, validating the usefulness of salt responsive candidate genes based these newly designed microsatellite primer pairs.ThesisItem Open Access Micropropagation and simple sequence length polymorphism analysis in Eggplant(DRPCAU, Pusa, 2020) Yadav, Kanchan; Anjani, KumariA study was conducted for micropropagation of the selected eggplant genotypes and to evaluate microsatellite based genetic variation among promising eggplant genotypes. The molecular data generated was used to elucidate the pattern and extent of differentiation and divergence at molecular level among these promising eggplant genotypes. The micropropagation was carried out using leaf as explant. MS media was chosen as the basal medium and five media with different combinations of hormones was used for culture. The leaves of the three selected genotypes were collected from Vegetable Farm, RPCAU, Bihar and cultured on all the selected media. Responses to tissue culture with regard to the frequencies of established aseptic cultures and their subsequent responses for swelling, callogenesis, caulogenesis and rhizogenesis were observed at regular intervals. The data was collected and subjected to statistical analysis and variance analysis. The best medium for swelling, callogenesis and caulogenesis response of cultured leaf explants was found to be M2 (MS + 2mg/l 2,4D + 0.01mg/l BAP). Among all genotypes, highest response of swelling and callogenesis was shown by genotype Laalgulab. GBL3 was found to have the best caulogenesis response than the other two genotypes. The molecular characterization of twelve genotypes was done using seven SSR markers. The microsatellite marker based amplification of 12 genotypes detected 56 alleles, out of which 23 alleles were found to be unique. Among all the primer pairs, primer pair EM114 was found to have the highest polymorphism per cent and PIC value. The highest value of discrimination coefficient was observed for the primer pair EM114. Estimates of similarity coefficients indicated the existence of considerable similarity among the genotypes. However, most of the genotypes had very low value of similarity coefficients, exhibiting the presence of high diversity among the twelve genotypes. The highest Dice similarity coefficient was observed for the genotypes Gol Banarasi and BB85. The cluster analysis differentiated the genotypes into different clusters. The similarity coefficient based dendrogram, grouped the genotypes into three clusters; cluster 1 included Gol Banarasi, BB85, Neelam Long, Laalgulab, PB6, Rajendra 2; cluster 2 included Bangal Bhatta, PS, GBL4, PB71, SMB 115 and cluster 3 included Arka Keshav. The molecular study showed the presence of diversity among the twelve genotypes of eggplant used in the present study.ThesisItem Open Access Effect of heat stress on wheat tillering and its molecular characterization(DRPCAU, Pusa, 2020) Vivek, Kommu; Kumar, RajeevTillering is one of the most important agronomic traits in cereal crops like wheat. It determines the number of spike and panicles per plant, thus affects the biomass and grain yield. It is very sensitive to temperature and often gets affected by the heat stress resulting in low yield in the crop. In the current study, 40 doubled haploid (DH) lines were sown under normal and late sown conditions to assess the effect of temperature on tillering and grain yield. Under normal sown conditions, the average tiller number was 154.6 tiller per meter with grain yield of 170.7g. During the period, the average temperature between the sowing to the jointing stage was 24.6℃. While, during the tillering (21DAS to 45DAS), the average temperature was recorded as 20.3℃. The DH lines JR 7, JR 56, JR 75, JR 80, JR 117, JR 118, JR 142, JR 143, JR 159, PBW343 fell in the category of high tillering under normal sown unstressed growing condition. Under the late sown condition, the average tiller number was 136.05 tiller per meter, with the average grain yield of 96.36 g. The average temperature between the sowing to the jointing stage of the late sown was 22.8℃. While during the tillering (21DAS to 45DAS), the average temperature was recorded as 21.9℃. The DH lines JR 2, JR 56, JR 104, JR 118, JR 119, JR 142, JR 143, JR 148, JR 168, KSG 1186 exhibited high tillering during the late sown (stressed) condition. Analysis of variance indicated a significant difference among the doubled haploid lines for the character under consideration for two different dates of sowing. The available sequence information of “tin” and monoculm genes, related to tillering in the plant, was used, and three gene-specific microsatellite primer pairs were designed with the help of appropriate bioinformatics tools. The primer pairs were used to characterize 40 DH lines under study. Reproducible amplification was achieved. Polymorphic information content of these 3 microsatellite primers ranged from 0.8 to 0.9, with an average value of 0.89. A total of 36 alleles were detected in the studied lines, out of which 7 were unique, and 29 were shared alleles. The genes specific designed microsatellite primers revealed ample genetic divergence at the molecular level amongst the wheat genotypes. Pair-wise Similarity Coefficient ranged from 0 to 1. Cluster analysis grouped the DH lines understudy into nine different clusters at phenone level 10 (coefficient 0.1).ThesisItem Open Access Amplification profiling of grain zinc accumulation related candidate genes in rice using TRAP markers(DRPCAU, Pusa, 2020) Lalaso, Abhang Sachin; Sharma, V.K.The present study was undertaken to examine the molecular level genetic polymorphism in relation to grain zinc concentration among 18 genotypes of rice using 12 candidate gene specific TRAP markers. The purpose of the present investigation was to evaluate the extent of genetic variability and diversity in respect of grain zinc concentration among these rice genotypes. Differential amplification profile was ascertained amongst the genotypes subjected to molecular characterization by employing 12 TRAP markers. Altogether 141 polymorphic bands representing easily recognizable different types of amplified products were categorized into 29 unique products and 112 shared products. Polymorphism information content of 12 TRAP markers diverged from 0.437 to 0.776 around an average measure of 0.611 across the markers. The combination of fixed forward primers OsNRAMP7K, OsZIP3K, OsZIP4a and OsZIP5K with an arbitrary reverse primer ME02 generated the lowest number of polymorphic products. Furthermore, the forward primer OsZIP7K in combination with the same arbitrary reverse primer yielded the highest number of polymorphic products. The forward primers OsZIP1-1, OsZIP7K, OsZIP4a, OsZIP5K, OsZIP4K, OsYSL14K and OsZIP5-3 along with ME02 yielded remarkably larger allelic size difference in descending order of magnitude. Numerical variation in polymorphism information content of the markers reflected their allelic diversity and distribution frequency among the genotypes, which diverged from 0.437 for the combination of primer OsZIP5-3+ME02 to 0.776 for the combination of primer OsZIP7-2+ME02 with an average of 0.611 across all the primer pair combinations. The fixed forward primers OsZIP7-2, OsZIP1-1 and OsZIP4K in combination with ME02 seemed to be extremely polymorphic and informative, whereas the remaining nine primer pair combinations were found less polymorphic. Similarity coefficients measuring the genetic similarities on the basis of the total count of shared polymorphic amplified products among pair-wise combinations of the genotypes revealed sizeable molecular level genetic variation among the genotypes. Experimental results clearly reflected the existence of plentiful diverseness at the molecular level among the 18 genotypes under evaluation in the present study. Hierarchical classification pattern based on target region amplification polymorphism related genetic similarity among rice genotypes was almost in complete agreement with principal coordinate analysis based two-dimensional dispersion pattern of molecular genetic profiles of rice genotypes. Using both the modules, the genotypes were conveniently grouped into four considerably discriminated multi-genotypic groups highly consistent with their grain zinc concentration. Candidate gene markers based analysis appeared remarkably effective in differentiation of rice genotypes in relation to their grain zinc concentration. Hence, these markers can be efficaciously and expeditiously employed for grain zinc concentration associated distinction of rice genotypes and extraction of rice genotypes for their utilization as parental lines during genetic enhancement in respect of grain zinc biofortification. Significance of zinc transporters based target region amplification polymorphism markers for differentiation and divergence analysis was established in relation to grain zinc concentration in rice genotypes.ThesisItem Open Access Characterization of little millet genotypes using morphological traits and ISSR markers(DRPCAU, Pusa, 2020) C, Shankar; Anjani, KumariThe present investigation entitled ‘‘Characterization of Little Millet Genotypes Using Morphological Traits and ISSR Markers’’ was undertaken with the objective of evaluating little millet genotypes for its morphological characters and molecular characterization based diversity analysis using ISSR markers. Altogether 18 little millet genotypes were obtained from the Hill Millet Research Station, Gujarat. During kharif season 2020, the genotypes were evaluated at TCA, Dholi, Agricultural farm of Dr. RPCAU, Pusa, Bihar in RBD design with three replication. The genotypes were evaluated for its four morphological traits viz., plant height (cm), total number of tillers per plant, length of flag leaf (cm) and width of flag leaf (cm). Based on the ANOVA analysis, the maximum mean sum of square was recorded for flag leaf length (24.249) followed by plant height (10.734) between the replication. The mean sum of squares value of plant height within the genotypes was highest (832.535) followed by flag leaf length (170.175) within the genotypes. The mean sum of square values of all four trait within the genotypes are highly significant at table value P = 0.05 whereas the mean sum of square within the replication are not significant. The ANOVA analysis revealed the presence of significant amount of variability for plant height, length of flag leaf, number of tillers per plant and width of flag leaf. Among the studied quantitative traits, plant height and length of flag leaf were the most contributing traits towards total diversity. The mean values of four quantitative traits were subjected to descriptive statistical analysis. Plant height ranged from 59.94 to 152.4 with the highest mean value of 102.95 followed by length of flag leaf ranged from 35.3 to 78.48 with the mean value of 54.73. The plant height (92.46) followed by length of flag leaf (43.18) recorded wide range which indicate the extent of variability for the two phenotypic traits. A total of 16 ISSR primers were used to identify the polymorphism between the 18 little millet genotypes. All the primers produced the polymorphic amplification product in 18 genotypes except HB 10, UBC 814 and UBC 817. The amplification product size ranged from 354 to 2315 bp. Total number of alleles detected were 370 including 162 unique alleles and 208 shared alleles. The polymorphism per cent ranged from 18.18% for the primer UBC 864 to 78.12% for the primer UBC 807 with the average of 38.66. The 16 ISSR primers included in the study had high polymorphism per cent except the primer UBC 816. The Polymorphism Information Content value ranged from 0.897 for the primer UBC 864 to 0.989 for the primer UBC 807 with the average of 0.969. Out of 16 ISSR primers, the primer UBC 807 was determined to the best in its effectiveness to detect the genetic diversity among the 18 little millet genotypes followed by HB 10 and UBC 814. The Dice’s similarity coefficient value ranged from 0.0208 to 0.5055. The similarity value was highest between the genotypes WV-168 and WV-119 was 0.5055. The higher the similarity coefficient values, greater will be the similarity between the pairs. The similarity coefficient value for the most of the genotype pairs was less than 0.4 among the group which indicates the presence of high amount of genetic variability between the genotypes of the little millet. The 18 genotypes of little milletswere grouped into 5 clusters at 50 phenon level by UPGMA dendrogram. The cluster I was formed by four genotypes (WV-155, WV-146, WV-168 and WV-119). The cluster II was formed by four genotypes (JK-18, TNPS-173, GLM-203 and WV-159). The cluster III was formed by four genotypes (TNPS-171, WV-108, WV-110 and WV-122). The cluster VI was formed by two genotypes (TNPS-167 and WV-167). The cluster V was formed by four genotypes (WV- 156, WV-124, WV-115 and WV-164). The cluster analysis revealed the presence of diversification among the little millet genotypes. Thus ISSR markers are highly efficient for the assessment of genetic relationship at molecular level which can be utilized in crop development programme.ThesisItem Open Access Identification and characterization of important mineral transporters in rice (Oryza spp.)(DRPCAU, Pusa, 2020) Kothari, Shubham; Sharma, Vinay KumarA study involving genome wide analysis of genomic and peptide sequences of eleven species and sub-species of rice (Oryza spp.) was undertaken for identification and characterization of Calcium and Cadmium mineral transporters. Retrieval and analysis of genomic and peptide sequences was carried out with the help of different methodologies and tools including SMART, MeV, FASTA, Clustal, Ortho-Venn, MEME suite, HAMMER and Cygwin64. Only 577 genes were identified as true transporters through annotation program among 797 calcium transporter gene sequences isolated from the database. Among these transporter genes, 452 proteins were orthologues, whereas, 25 genes were species specific. The maximum (63) and minimum (44) number of calcium transporters were identified in Oryza sativa indica and Oryza sativa japonica, respectively. Computational analysis of cadmium transporters revealed 885 genes for true transporters among 893 gene ids. Across all the species and sub-species, 585 proteins were orthologues, whereas 32 genes were species specific. The maximum (95) and minimum (67) number of cadmium transporters were identified in Oryza sativa indica and Oryza longistaminata, respectively. Phylogenetic analysis grouped the identified 452 calcium transporters in to five major super families, namely, PM_ATPase, CT_ATPase, CZT/CuT, PLT_ATPase, CACC/CPCC with majority of the mineral transporters belonging CT_ATPase family and a smaller number of transporters belonged to CZT/CuT_HMA family. Similarly, cadmium transporters were grouped into six major super families, namely, ABC type transporter; ATPase_type transporter; Sulphate transporter; Protein NRT1/ PTR family; Metal transporter NRAMP5 and MACPF domain-containing protein CAD1. Most of these mineral transporters belonged to ATPase_Type family and lowest number was found in NRAMP5 and CAD1 family. For calcium transporters, orthologous determination study revealed the maximum number of orthologous protein count in Oryza sativa indica (45), while its minimum number in Oryza sativa japonica (35) for calcium transporters. Similarly, for cadmium transporters, maximum number of orthologous protein count was found in three species (58), namely, Oryza barthii, Oryza sativa indica and Oryza punctata, whereas its minimum was recorded in Oryza sativa japonica (44). Motif analysis revealed the presence of some conserved domain ids in all the species and sub-species of rice under investigation in this study. Family distribution pattern of highest clustered protein indicated highest predominance of CT-ATPase (Calcium Transporter ATPase Transporter) sub-family of transporter proteins in the case of calcium. Similarly, ATPase-type transporter (ATPase dependent Transporter) sub-family was abundant with highest number of transporter sequences in the case of cadmium. Computational results provided the basis to infer gene expression in shoot tissues in the case of calcium, while gene expression seemed to occur only in seed without any expression in shoot/root/leaves in the case of cadmium. Furthermore, high level of expression of predicted calcium transporter genes into seeds and its parts rather than the shoot portion suggested that these proteins might be involved in calcium accumulation in seeds, embryo and endosperm parts. Nine gene ids (Os02t0176700-00, Os02t0196600-01, Os03t0203700-01, Os03t0281600-01, Os03t0689300-01, Os04t0605500-01, Os04t0656100-01, Os07t0191200-01, Os12t0638700-01) had sequence similarity in the case of both the minerals, reflecting that these genes are responsible for the transportation of both calcium and cadmium in Oryza sativa japonica. So, these may be considered as calcium-cadmium co-transporters. Experimental results showed that many of the sequences are distributed in the genome of different species and sub-species of rice. Significant evidences were generated for exploring the expression and function of these calcium and cadmium transporters under different environmental conditions at different tissues in plant to understand the molecular basis of mineral accumulation in developing grains.ThesisItem Open Access Molecular fingerprinting of locally adapted landraces and improved varieties of rice using ISSR markers(DRPCAU, Pusa, 2020) Kumar, Ravi; Bhutia, K.L.The level of genetic variability present in the population primarily depends on genetic progress. The molecular marker is a beneficial method for determining genetic variants and resolving the identities of cultivars. The goal of this research was to evaluate the diversity between 18 rice genotypes using 13 ISSR primers. Thirteen ISSR primers generated total of 81 amplified alleles with an average of 6.24 alleles per primer. Highest number of alleles i.e. 9 was obtained from the primer ISSR-834 and lowest 4 from ISSR-872 and out of 81 amplified alleles, five were rare alleles and eight were unique alleles. Amplified fragment size ranges from 148 bp to 2500 bp. Primer ISSR-880 showed very high PIC value (0.41) followed by four primers namely ISSR-872, ISSR-890, ISSR-823, ISSR-864 with high PIC values of 0.38, 0.32, 0.31, and 0.30, respectively, and rest of the primers showed medium PIC values ranging from 0.26 to 0.29. The highest value of EMR (7.11) was showed by the ISSR-834 followed by 7 in ISSR-890 and ISSR-864. The lowest value of EMR was observed in ISSR-888 and ISSR-846 with the value of 2.67 and 3.20, respectively and rest of the primers had EMR values ranging from 2.67 to 7.11. ISSR primers namely; ISSR-890, ISSR- 13 864, ISSR-834, ISSR-880 showed highest value (2.22, 2.10, 2.09 and 2.04) for marker index (MI) because these markers had higher value of either PIC or EMR or both (As MI is the product of PIC * EMR). ISSR-888 and ISSR-846 showed the lowest values (0.75 and 0.83) of MI and rest of the primers showed moderate value of MI ranging from 1.31 to 1.64. Resolving power of ISSR primers was highest in ISSR-890 (10.5) followed by ISSR-891 (9.55): ISSR-834 (8.78): and ISSR-841 (8.00). Primers with lowest RP values were ISSR-846 and ISSR-872 with RP of 2.89 and 2.44, respectively. Nei‟s Genetic Diversity (h) and Shannon Index (I) within the population of 18 genotypes yielded the value of 0.2961 and 0.4474, respectively, suggesting that the population of 18 rice genotypes is moderately diverse. Phylogenetic analysis and factorial analysis on the basis of Dice dissimilarity index grouped the 18 genotypes into 3 major cluster with cluster-I having the highest numbers of genotypes (eight genotypes) followed by cluster-II with six genotypes and cluster-III with four genotypes. The genetic Structure of 18 genotypes at K=2 (on the basis of 13 ISSR primers with 81 amplified loci) revealed that the 18 rice genotypes are the admixture of two ancestral components present in different combinations in 18 genotypes. The genetic structure explained by 81 amplified loci of 13 ISSR primers doesn‟t give the higher resolution of the actual genetic structure of 18 rice genotypes because the number of primers used is quite less and it is not known that how many chromosomes have been covered by these 81 amplified loci. However, the 13 ISSR primers used were sufficient enough to differentiate between the 18 genotypes under study with many of the primers having very high PIC, EMR, MI and RP valuesThesisItem Open Access Genetic analysis of maize inbreds and hybrids using biochemical and microsatellite markers(DRPCAU, Pusa, 2020) Suman, Sandeep Kumar; Kumar, MithileshCharacterization of inbred lines and hybrids is crucial for release of hybrid and genetic purity analysis. Hence, field evaluation was done at AICRP, Maize, TCA, Dholi farm and lab experiment was done at Molecular Biology Lab, Department of Plant Breeding and Genetics, RPCAU, Pusa for characterization of parental lines and hybrids and genetic purity analysis through biochemical and microsatellite marker. Morphological genetic diversity analysis between thirteen inbreds of maize were done using Mahalanobis D2 statistics for ten characters viz. plant height, ear height, days to 50% tasseling, days to 50% silking, days to 50% Brown husk, ear length/Cob length, ear girth, number of kernels per row, number of kernel rows per cob, grain Yield per Plant. All the thirteen inbred lines under study were grouped in three clusters. Cluster I comprised of eleven inbred lines G18, CLQRC, WLS, HK1, CML490, S8200, S8481, R6429, R4093, VL111 and CLQR. Cluster II and III were found monogenotypic and consisted of CLQ25 and CG18 respectively. The inter-cluster distance between cluster II and cluster III was found maximum 6.67, while lowest inter cluster distance value was found between 4.62 cluster I and cluster II. The maximum value of genetic distance 4.89 was estimated between inbred line WLS and G18, while minimum genetic distance 2.38 was found in between VL111 and S8481. Name of the Student : SANDEEP KUMAR SUMAN Registration No. : D/AB/130/2014-2015 Major Advisor : Dr. MITHILESH KUMAR Degree to be Awarded : Ph.D. (Agricultural Biotechnology) Department : Agricultural Biotechnology and Molecular Biology Major Subject : Agricultural Biotechnology Minor Subject : Plant Breeding and Genetics Year : 2020 Total pages of the Thesis : 144 + xviii (Bibliography) + xxii (Apppendices) Title of Thesis : “Genetic analysis of maize inbreds and hybrids using biochemical and microsatellite markers” The result showed great variability was found in genetic makeup of the inbred lines. Cluster I accommodated eleven inbred lines having maximum mean value for number of kernels row per cob and number of kernels per row. While, minimum value was observed for ear height and plant height. Maximum mean value for grain yield per plant and ear girth was found for inbred line CLQ25 that is present in cluster II. The maximum contribution 26.63% was found for the trait grain yield in manifestation of genetic divergence followed by ear girth (28.47%), ear length (19.23%), number of kernel per row (11.50), number of kernels row per cob (10.04). Out of thirty crosses, eleven crosses manifest positive significant heterobeltiosis for the trait grain yield per plant. Two crosses, CLQ25 × CLQR and WLS × R4093 belongs to high divergent class. It could be deduced that genetic diversity can be used as an authentic parameter to predict heterosis in hybrids. The parental lines and hybrids were characterized with SDS-PAGE using total salt soluble seed proteins. Total salt soluble banding profile identified one hybrid G18 x VL111. The presence of 53 KDa, MW band specific for female parent G18 and 65 KDa, MW band specific for female inbred line confirmed the hybridity and purity of hybrid G18 x VL111. The 27 microsatellite primer amplified a total of 28 loci with an average of 4 alleles per locus. The number of alleles varied from 2 to 8 in case of umc1222 and umc1545 or umc1265 or umc1963 or phi084, respectively. The range of amplification varying from 72-88 in case of umc1327, while in case of phi0116 amplification ranged from 293-345. A total of 112 alleles were found in which 33 were unique and rest 79 were found shared alleles in 13 inbred lines. Based on complementary banding pattern between hybrids and their parents, the six microsatellite markers viz. umc1222, umc1161, umc1196, umc1367, umc1403 and nc133 were found effective marker to distinguish F1 hybrids from its parental lines. These markers were utilized to analyse genetic purity of inbred lines and hybrids.ThesisItem Open Access Genome profiling for differential grain iron concentration in rice using candidate gene based microsatellite markers(DRPCAU, Pusa, 2020) Namdev, Kale Anil; Sharma, V.K.A study was performed to evaluate the genetic polymorphism by employing candidate gene related microsatellite specific designed primers for validation of their significance in discrimination of rice genotypes in relation to grain iron concentration. Utilizing a set of 12 microsatellite primers targeting 6 candidate genes related to five chromosomes, amplification of targeted region in genomic template of 18 rice genotypes was achieved. Statistical measures and parameters employed to derive inferences included polymorphism information content, polymorphic per cent and similarity coefficient. Newly designed microsatellite primers directed amplification was successfully achieved. Among the 6 candidate genes, single microsatellite sequence for OsYSL4 and OsYSL6, two microsatellite sequences for OsMTP1, OsYSL1 and OsNAS3 and four microsatellite sequences for OsNAS2 were investigated to analyze the molecular level genetic polymorphism amongst the entries. Altogether 12 microsatellites were investigated in which one, five and six microsatellites had tetra-nucleotide, di-nucleotide and tri-nucleotide repeat motif, respectively. Differential migration of amplicons resulted in appearance of bands representing amplification products localized at different positions on the gel, reflecting the variation in product length derived by primer directed amplification of genomic templates. Altogether 46 shared and 16 unique alleles with a mean value of 5.16 alleles per primer pair were recognized among 62 allelic variants. Null allele was also noticed in the specific combinations between genotypes and primers. The polymorphic information content of panel of primers exhibited appreciably higher order of variation with the value ranging from 0.685 (OsYSL1f) to 0.870 (OsYSL1a) and mean value of 0.756 per primer pair. Differential amplification pattern reflecting genomic level polymorphism between genotypes under evaluation provided a basis for deducting that length variation of repeat sequences observed within candidate genes may be influential in differential accumulation of iron in grains of rice. Sizable variation was clearly recognized and reflected by deviations of repeat sequence length in the genomic region flanked by forward and reverse primers. Similarity coefficient indices based on proportional measures of shared alleles showed maximum similarity (0.727) between ERG-10 (Rajendra Bhagwati) and ERG-09 (BPT5204 Sub1). Similarity coefficient based hierarchical cluster analysis resulted in discrimination of 18 rice genotypes into three major genotypic groups and subsequently into five clusters. Genotypic discrimination was highly effective in relation to grain iron concentration of rice genotypes. Appreciably greater degree of molecular level genetic distinction and diverseness was exposed between genotypes characterized by molecular profiling. Hierarchical pattern of classification based on similarity coefficient matrix of pair-wise genotypic combinations was in extremely good agreement with two-dimensional distribution pattern revealed by principal coordinate analysis of genetic profiles of entries. Using these two approaches of genotypic discrimination based on analysis of genic microsatellite-specific markers derived from candidate genes made it possible to differentiate and classify the entries with appreciably greater level of consistency in relation to their grain iron concentration. Experimental results amply emphasized that the panel of primers employed during experimentation was useful and adequate to discriminate the rice entries in relation to grain iron concentration. These markers can therefore be used effectively and efficiently to discriminate the genotypes of rice and to facilitate selection of diverse parents during genetic enhancement in relation to grain iron biofortification.
