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Chaudhary Charan Singh Haryana Agricultural University, Hisar
Chaudhary Charan Singh Haryana Agricultural University popularly known as HAU, is one of Asia's biggest agricultural universities, located at Hisar in the Indian state of Haryana. It is named after India's seventh Prime Minister, Chaudhary Charan Singh. It is a leader in agricultural research in India and contributed significantly to Green Revolution and White Revolution in India in the 1960s and 70s. It has a very large campus and has several research centres throughout the state. It won the Indian Council of Agricultural Research's Award for the Best Institute in 1997. HAU was initially a campus of Punjab Agricultural University, Ludhiana. After the formation of Haryana in 1966, it became an autonomous institution on February 2, 1970 through a Presidential Ordinance, later ratified as Haryana and Punjab Agricultural Universities Act, 1970, passed by the Lok Sabha on March 29, 1970. A. L. Fletcher, the first Vice-Chancellor of the university, was instrumental in its initial growth.
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ThesisItem Open Access Linkage mapping of quantitative trait loci for root morphology and selection of desirable genotypes from aerobic x lowland indica crosses in rice (Oryza sativa L.)(CCSHAU, 2014) Kharb, Anju; Jain, Rajinder KumarIdentification of stable QTL for traits promoting aerobic adaptation using molecular markers can greatly enhance the efficacy of breeding programs to develop water-efficient, high yielding, aerobic rice varieties. Filial (F2 and F3) and backcross ( BC2F2 and BC1F2) populations derived from the crosses between high-yielding low-land (HKR47) and aerobic (MAS26 and MASARB25) indica rice varieties, displayed large variation for various physio-morphological traits including grain yield per plant and root traits (in case of net house evaluation). Phenotypic correlation analysis showed positive correlation (r = 0.279 and 0.351) between grain yield per plant and root length in both F2 and F3 population and between grain yield per plant and fresh & dry root weight (r = 0.232 and 0.269) in F2 population. Under aerobic field conditions, Grain yield per plant showed significant positive correlation with plant height, effective number of tillers/plant and panicle length in all the four populations. A total of 803 SSR markers, distributed on 12 rice chromosomes, were analyzed for polymorphism in two parental rice genotypes (HKR 47 and MAS 26); of these 125 (about 16%) displayed polymorphism. SSR fingerprint databases of three HKR47 x MAS26 F2 and F3 populations (94 F2 in net house, 42 F3 in net house and 52 F3 plants in field) were prepared using 125, 88 and 88 polymorphic SSR markers, respectively. NTSYS-pc UPGMA tree cluster analysis and two-dimensional PCA scaling showed scattering of the F2 and F3 populations between the two distinct parental genotypes; the populations were inclined towards MAS26. Composite interval mapping (CIM) analysis by Win QTL cartographer 2.5 revealed a total of six QTLs (qPN8.1, qPN8.2, qPN8.3, qTGW8.1, qYPP8.1 and qRL8.1) in HKR47 x MAS26 F2 population, grown in the net house on chromosome 8 (within a region of 24.9 cM) which individually explained 13.7-27.3% of the phenotypic variation. In case of HKR47 x MAS26 F3 population, based on field data and net house data, two (qTGW6.1 and qLB8.1) and six QTL (qPH3.1, qLB8.1, qTGW6.1, qTGW11.1, qRT8.1 and qRT8.2) were mapped, respectively. As many as 20 F3, 6 BC2F2 and 5 BC1F2 promising plants having higher grain yield and/or better root length/biomass were selected. Data on monitoring of putative QTL identified showed that out of twenty selected plants, four plants had 13 QTL, three plants had 12 QTL and two plants had 11 QTL in homozygous or heterozygous condition.ThesisItem Open Access Molecular Dissection of AGPase to Improve Wheat Productivity(CCSHAU, 2014) Batra, Ritu; Sikka, V.K.The present investigation was conducted towards understanding role of ADP-glucose pyrophosphorylase (AGPase) in developing grains’ starch biosynthesis among diverse wheat lines including disomic chromosome substitution (DCS) lines of C-591 in the background of Chinese Spring, dwarf wheat varieties. isogenic lines and for WH-711 X WH 542. The experiment was laid out in completely randomized block design with three replications. The traits like grain growth rate (GGR) and AGPase enzyme activity were also measured for developing grains at 10th, 20th and 30th days after anthesis (DAA). Metabolites like total sugars, reducing sugars, non-reducing sugars, total starch, amylose and amylopectin were also quantified using wheat flour of the test lines. The recorded data was subjected to appropriate statistical analysis to arrive at inferences on the experiments in the study. Critical analysis of the data led us to conclude that determinants for AGPase expression are present across the genome over multiple chromosomes which are active at particular grain filling stages like during first half, chromosome 1B and 2D played major role whereas second half was mainly controlled by chromosome 6A and 5B. Hybridization efforts have improved grain attributes and yield components of WH-542 with ~28.6 % and 42 % enhancement in grain wt. per spike and yield per plant respectively. Maximum amylose (%) was observed in DCS line 2D which can be used to produce functional wheat flour. Grain yield was found to be significantly associated with grains/spike and plant biomass in case of DCS lines. Partitioning the variation by path analysis indicates grains per spike, though had less direct effect but its impact on grain yield through indirect effect via tiller no./plant and AGPase activity at 20th DAA was high suggesting their importance in yield improvement efforts. AGPase activity analysis suggests major contribution of chromosome 2D towards yield in a specific temperature window of 22±1.5oC during initial stages of grain filling which can be used as a screening tool for terminal heat tolerant genotypes.ThesisItem Open Access Selection of high-yielding iron-rich PAU201/Palman579 segregating rice (Oryza sativa L.) lines using conventional and molecular marker techniques(CCSHAU, 2014) Naveen Kumar; Chowdhury, V.K.Molecular markers provide novel tools for linkage mapping of QTLs of target traits and can greatly enhance the efficacy of breeding programs to improve mineral (iron and zinc) density in rice. F3, F4, BC1F2 and BC1F3 populations derived from the cross between high-yielding (PAU201) and iron-rich (Palman 579) indica rice varieties displayed large variation for various physio-morphological traits including grain yield per plant and mineral (iron and zinc) contents. Iron and zinc content varied from 0.9- 149.9 and 0-143.1 μg/g respectively in all the four populations (F3, F4, BC1F2 and BC1F3). Transgressive segregation for grain iron content was noticed in F3 population with one of the plants having exceptionally higher iron (746.8 μg/g) content. Phenotypic correlation analysis showed positive correlation (r=0.281) between grain iron content and zinc content in BC1F2 population but not in F3, F4 and BC1F3 populations. Grain yield per plant showed significant positive correlation with plant height and effective number of tillers/plant in all the four populations. A DNA fingerprint database of 33 PAU201 × Palman 579 F4 plants was prepared using 61 polymorphic SSR markers distributed on the entire genome of rice. NTSYS-pc UPGMA tree cluster analysis and two-dimensional PCA scaling showed scattering of the F4 population between the two distinct parental genotypes; the population was inclined towards Palman 579. The distribution of PAU201 and Palman 579 specific alleles among the PAU201 × Palman 579 F4 plants were determined and SSR data was used to identify QTLs for grain mineral content and various agronomical traits. A total of 128 alleles were identified in 33 PAU201 x Palman 579 F4 plants and three new recombinant alleles (different that those in parent rice varieties) were identified. Composite interval mapping (CIM) analysis by Win QTL cartographer 2.5 revealed a total of six QTLs for mineral content (five for Fe and one for Zn) in rice grains on chromosome 5, 6, 7 and 9; and sixteen QTLs for various agronomical traits (plant height, tillers per plant, yield per plant, Grains per panicle and 1000-grain weight). As many as 33 F4 and 25 BC1F3 promising plants having higher grain yield as well as mineral content have been selected for further progeny analysis.ThesisItem Open Access Marker assisted selection for drought tolerance in pearl millet (Pennisetum glaucum (L.) R. Br.)(CCSHAU, 2014) Asha Rani; Yadav, Ram C.Pearl millet (Pennisetum glaucum (L.) R. Br.) is the staple cereal of millions of poor rural families in the hottest and driest areas of the tropics and subtropics. Terminal drought stress is a regular occurrence and serious environmental constraint in these regions, making stress tolerance an essential attribute of new pearl millet cultivars. Marker assisted selection is an important molecular tool for improving yield and yield stability of the pearl millet. A number of genomic regions were found to be associated with drought tolerance in terms of both grain yield and its components. A major QTL associated with the drought tolerance of grain yield mapped on linkage group 2 and explained up to 32% of the phenotypic variation. Another QTL were observed on LG 5 with phenotypic variation of 14.8%. During the present investigation, HHB 226 pearl millet hybrid was improved for drought tolerance by marker assisted selection. For this, two crosses were made i.e. HBL 11 x PRLT 2 and HBL 11 x 863 B. PRLT 2 and 863 B were used as donor parent and HBL 11 (male parent of HHB 226) was used as recurrent parent. Eleven SSR primers specific to drought tolerance from LG2 and 5 were used for polymorphism analysis between recurrent and donor parent. Three SSR primers from LG 2 (PSMP2066, PSMP2077, and PSMP2059) and one from LG 5 (PSMP2078) were found polymorphic in both the crosses which were used for further screening. Recurrent and donor parents were crossed to raise the F1 generation. Selected plants were backcrossed with HBL11 three times and in each generation positive plants were selected. Finally, four plants were selected in BC3F1 generation in both crosses which were then used for background selection. 25 and 32 background primers were found polymorphic out of 64 primers in HBL 11 x PRLT 2 and HBL 11 x 863 B cross respectively. About 70 and 80% of the recurrent parent genome was found to be transferred in most of the selected plants in 1st and 2nd cross respectively till BC3F1 generation. During the drought screening test in BC2F1 generation in 1st cross, RWC and 1000 seed weight were found to be enhanced with 27.84% and 15% respectively, in drought tolerant BC2F1 generation plants than drought sensitive. In 2nd cross, RWC and 1000 seed weight were increased with 33.6% and 28% respectively, in drought tolerant plants as compared to drought sensitive.ThesisItem Open Access Expression profiling of soluble starch synthase I gene during grain development for thermotolerance in wheat (Triticum aestivum L. em. Thell)(CCSHAU, 2014) Gupta, Meenu; Dhillon, SantoshThe four genotypes of wheat i.e. two thermotolerant (WH 730 and WH 1021) and two thermosensitive (WH 147 and WH 711) were evaluated under two different sowing conditions (normal and late sown) to assess the effect of high temperature (HT) stress on grain yield traits and expression of soluble starch synthase I gene. Gene expression was studied by real time PCR analysis at 7 stages (2, 5, 8, 10, 15, 20 and 25 DAA) during grain development in developing grains and flag leaf. High temperature stress negatively affected all yield traits like number of tillers per plant, grain number per plant, thousand grain weight, biomass and grain yield per plant. Decreased grain filling duration under HT stress was not compensated by increased grain filling rate. The effect was more profound in case of thermosensitive genotypes. The order of thermotolerance based on heat susceptibility index (HSI) is: WH 730 (0.65) > WH1021 (0.79) > WH 711 (1.13) > WH 147 (1.52). In developing grains, temporal pattern of SSSI gene expression was altered and expression peaked earlier under LS conditions (15 DAA) than under NS conditions (at 20 DAA) in all genotypes studied. In flag leaf, peak expression under NS conditions was observed at 15 DAA in all genotypes while under LS conditions, expression peaked at 15 DAA in thermotolerant genotypes and at 10 DAA in thermosensitive genotypes. At late stages during grain development, SSSI gene expression fell more abruptly and to a higher level in case of thermosensitive genotypes in grains as well as flag leaf. Partial SSSI cDNA was sequenced from WH 730 (1916 bp), WH 1021 (1924 bp), WH 147 (1925 bp) and WH 711 (1924 bp) genotypes. Phylogram generated using different starch synthases isoforms clustered SSSI, SSSII, SSSIII and SSSIV into separate groups. WH 730 showed maximum tolerance to HT stress while WH 147 was least thermotolerant, based on the overall picture of physiological traits, yield traits and gene expression analysis studies.ThesisItem Open Access Molecular and physiological studies to identify traits enhancing rice cultivars adaptation to aerobic conditions for sustainable rice production in water short situations(CCSHAU, 2013) Sandhu, Nitika; Jain, Rajinder KumarIn the present investigation experiments were conducted to: identify QTL conferring a potential yield advantage under direct seeded conditions; identify root traits having a positive correlation with grain yield under direct seeded conditions; identify genotypes with high levels of root growth plasticity in multiple conditions (upland, lowland, induced by drought or low P) and to identify a positive correlation of root plasticity with grain yield; BSA approach was applied to identify QTL showing consistent effects in the background of two popular high-yielding varieties, IR64 and MTU1010, under upland reproductive-stage drought stress; identify QTL with large and consistent effects for traits thought to be beneficial for direct-seeded rice: seedling emergence, early vegetative vigour, root morphology, nutrient uptake, and grain yield. We hypothesized that the some of the seedling-stage traits investigated might have a strong enough effect on plant growth to be correlated with grain yield at harvest. Our study identified a total of 35 QTL associated with 14 traits on chromosomes 1, 2, 5, 6, 8, 9, and 11 in MASARB25 × Pusa Basmati 1460 population and 14 QTL associated with nine traits on chromosomes 1, 2, 8, 9, 10, 11, and 12 in HKR47 × MAS26-derived population. These identified QTL included three large-effect stable QTL for increased yield under aerobic conditions and QTL for several root-related traits likely to increase water and nutrient uptake under aerobic conditions. A number of plants were identified with higher root length and dry biomass, yield per plant, length-breadth ratio, and with Pusa Basmati 1460 specific alleles in homozygous or heterozygous condition at the BAD2 locus; these lines shall serve as novel materials for the selection of stable aerobic Basmati rice varieties. A good correlation was observed between root growth/plasticity in lysimeter, field and rhizoscope study, between water uptake in lysimeter and leaf water status in field. Consistently high plasticity was observed across different treatment and environment. Three large and consistent-effect QTL, qDTY1.1, qDTY2.2, and qDTY2.3 for GY were identified under reproductive-stage drought stress in Kali Aus/2*MTU1010 and Kali Aus/2*IR64 populations. A total of 28 QTL associated with 22 traits and 20 QTL associated with 12 traits were mapped in Aus276/3*IR64 and Aus276/3*MTU1010 population, respectively. The QTL qGY6.1, qGY10.1, qGY1.1 and qEVV9.1 were found to be effective in both Aus276/3*IR64 and Aus276/3*MTU1010 populations under wide range of conditions; the QTL for nutrient uptake were located on chromosome 5, co-located with qGR5.2 and qRHD5.1. The positive interaction between the genomic regions for root traits and nutrient uptake depicted that these interacting loci should also be considered when introgressing QTL to develop rice for dry direct seeded conditions. Co-localization of QTL for yield, EVV, and root traits indicates that the identified QTL may be immediately exploited in MAB to develop novel high-yielding direct seeded rice varieties.ThesisItem Open Access Development and characterization of Bt pigeonpea (Cajanus cajan L. Millspaugh) plants for resistance against Helicoverpa armigera (Hübner)(CCSHAU, 2014) Jain, Meenakshi; Kharb, PushpaPigeonpea [Cajanus cajan (L) Millsp.] is an important protein rich legume crop grown in many semi arid tropical regions of the world. Development of transgenics in pigeonpea remains dogged by poor plant regeneration in vitro from transformed tissue and low frequency transformation protocols. Transgenic pigeon pea plants of variety Manak (H 77-216) expressing cry1Ac gene were generated by tissue culture independent, rapid and efficient, transformation method using Agrobacterium tumefaciens strain LBA4404, harboring pBin-35Scry1Ac (Kharb et al., 2012). Putative transformants were screened at early stage by subjecting small leaf pieces to PCR amplification using plant direct PCR kit and transformation frequency of 45.0 % was obtained. Quantitative ELISA of 60 days old T0, transgenic plants showed expression of Cry protein ranging from 153.5ng/g – 562.5ng/g FW of leaf. Of the 374 seeds harvested from all the T0 plants, 324 were sown to obtain progeny (T1) plants and 233 of these were randomly analyzed to determine their transgenic nature. Out of the 233 T1 plants 125 amplified cry1Ac gene, substantiating the integration and transmission of the transgene in the transgenic pigeon pea plants. Gene integration was also checked in T2 plants by direct plant PCR The transformed (T1) plants genomic DNA showed single copy of cry1Ac gene integration by Southern hybridization and different transformation events were observed. Single copy for each transgenic event was also confirmed by real time PCR. Insect bioassay proved its usefulness over control plants for pod borer resistance. The protocol ensured generation of pigeon pea transgenic plants with considerable ease in a short time and offers immense potential for generating transgenic plants of difficult-to-regenerate pigeon pea.ThesisItem Open Access Phenotyping and microsatellite marker analysis of F2 and backcross populations derived from the crosses between aerobic and Basmati rice varieties(CCSHAU, 2014) Rajesh; Jain, R.K.Increasing water scarcity has threatened the cultivation of lowland Basmati varieties in northern states of India including Haryana. Marker assisted selection can greatly improve the efficacy of Basmati rice breeding programs. In the present investigation, experiments were conducted to evaluate two F2 populations and BC1F1 plants derived from crosses between Basmati (Improved Basmati 370, IB370) and aerobic rice varieties (MAS25 and MAS-ARB25), for various physio-morphological and root traits under aerobic conditions in field and nethouse. Both the segregating F2 populations displayed large variation for grain yield (3.9-40.9 and 2-43.6 g), root length (22-50 and 31-65 cm), root dry biomass (0.7-9.3 and 0.7-11.8 g), root thickness (17.4-30.4 and 10.2-32 mm) and grain length/breadth ratio (3.4-5.2 and 3.5-5.5), respectively. Phenotypic correlation analysis showed that root length was positively correlated (r=0.496) with root thickness in IB370 × MAS25 F2 population. Grain yield per plant showed significant positive correlation (r=0.250) with root thickness in IB370 × MAS-ARB25 F2 population. A DNA fingerprint database of two segregating F2 populations, IB370 × MAS25 and IB370 × MAS-ARB25 was prepared using 61 and 58 polymorphic SSR markers, respectively, distributed on the entire genome of rice. The distribution of IB370 and MAS25/MAS-ARB25 specific alleles among IB370 × MAS25 and IB370 × MAS-ARB25 derived F2 plants were determined and SSR markers data was used to identify QTL for various agronomic and root traits. Composite Interval Mapping (CIM) analysis by WinQTL cartographer 2.5 revealed a total of 13 QTL (six for root traits and seven for agronomical traits) on chromosome 1, 2, 3, 8 and 10 and a total of seven QTL (four for root traits and three for agronomical traits) on chromosomes 2, 4 and 5 in IB370 × MAS25 and IB370 × MAS-ARB25 derived F2 populations, respectively. Twenty promising F2 plants each from both the two crosses were selected on the basis of aroma, root length, L/B ratio and grain yield/ plant for further progeny analysis. The putative QTL identified in the two F2 populations were also monitored in the selected promising F2 plants; these QTL were present in homozygous or heterozygous state in high frequencies. DNA fingerprint database of 13 rice varieties was prepared using 129 polymorphic SSR markers, NTSYS-pc analysis and two-dimensional PCA scaling exhibited that 13 rice genotypes were interspersed in the two groups. Group I consisted of aerobic and indica, while group II had Basmati rice genotypes. Similarity coefficient data showed that Basmati rice variety is almost equally distant from aerobic indica (0.584) and lowland indica (0.549) rice genotypes.ThesisItem Open Access Transcript profiling of stress inducible MYB transcription factor and related genes in Indian mustard (Brassica juncea L. Czern. & Coss.)(CCSHAU, 2014) Aneja, Bharti; Yadav, Neelam R.Drought stress reduces the capacity of plants to take up water from soil, limiting plant growth and yield worldwide. The present investigation was carried out to study the expression of drought stress induced myb transcription factor genes (myb2, myb44, myb60) and related genes (hsp, pp2c and lhcb) in Brassica juncea cvs. RH0116 (drought tolerant) and RH8812 (drought sensitive) through semi-quantitative RT-PCR using actin gene as control. Brassica plants were raised in nethouse and subjected to drought stress by water withholding at flowering stage. We found that myb2, myb44, hsp and pp2c genes were upregulated and were expressed in both shoots and roots during drought stress. RH0116 showed higher accumulation of myb2, myb44 and pp2c transcripts as compared to RH8812. Whereas, myb60 and lhcb genes were downregulated during drought stress and their expression was limited only to shoots. The drought stress induced Atmyb44 homologues from RH8812 and RH0116 were cloned, sequenced and designated as BjMyb44-1 (872 bp) and BjMyb44-2 (862 bp) respectively. These had much similarity to other myb genes of Arabidopsis thaliana (84%) and Brassica rapa (94%). Multiple sequence alignment using CLUSTALW2 program with each other and with other sequences established homology. Seeds of both the genotypes were grown on MS medium with various levels of stress treatments using PEG 6000, Mannitol and ABA. Seed germination was arrested at initial stage only and reduction in seed germination was significant among treatments and no seedling growth was observed in case of ABA treatment. The average shoot and root lengths of seedlings decreased with the increasing dose of PEG & Mannitol in both the genotypes. The Brassica plants grown in vivo and two weeks old in vitro grown Brassica seedlings subjected to air drying stress treatments were used to assess the physiological parameters. The stressed plants showed lower Relative water content as compared to the control plants and the decline was higher in sensitive check. Osmotic potential was found to be more negative in case of stressed plants than control plants and decrease was relatively more in RH8812 as compared to RH0116. Electrolyte leakage increased in stressed plants as compared to control plants and RH8812 showed more increase than RH0116. Chlorophyll fluorescence study revealed that the value of ratio of variable fluorescence to minimum fluorescence (Fv/ Fm) was more in the control plants as compared to stressed plants, however, the increase was relatively more in RH8812 than RH0116.