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ThesisItem Open Access Evaluation of advanced wheat lines for stripe rust resistance and grain quality(CCSHAU, Hisar, 2019-08) Bhawna; Bhawna; Yadav, Neelam R.; Yadav, Neelam R.Biofortification of staple foods like wheat without any yield penalty is of utmost importance. Advanced lines introgressed with Gpc-B1 allele were used in the study. The Gpc-B1 allele co-segregated with Yr36 yellow rust resistance gene in all the advanced lines. All WH711-derived advanced wheat lines confirmed for the Yr36/Gpc-B1 allele. Seven advanced lines (WL-8-1, WL-8-2, WL-8-3, WL-8-5, WL-8-6, WL-8-7, WL-8-8) were confirmed for Yr18 while six advanced lines (WL-8-1, WL-8-2, WL-8- 6, WL-8-7, WL-8-8, WL-8-9) were positive for Yr17 gene. All five PBW343-derived advanced lines confirmed for the Yr36/Gpc-B1 allele presence. Three advanced lines (PL-8-2, PL-8-3, PL-8-4) displayed the presence of the Yr18 gene. Two advanced lines (Pl-8-3, PL-8-5) confirmed presence of the Yr17 gene. All the advanced wheat lines derived from WH711 as well as from PBW343 displayed very high levels of resistance against yellow rust and better agronomic characteristics than the parents. Positive correlation was observed in yield and yield related traits. The hectoliter weights of PBW343-derived lines indicated their superiority in terms of good flour recovery than PBW343 and comparable to the checks. WH711- and PBW343- derived advanced wheat lines show better gluten quality and high quantity in sedimentation test. Gpc-B1 allele introgression provided high GPC in WH711 derived advanced wheat lines. Iron and zinc contents of both WH711-derived and PBW343-derived advanced wheat lines were at par when compared to either parent and checks, indicating successful bio fortification of both micronutrients. Two of the WH711-derived lines are being further tested in IPPSN/station trials.ThesisItem Open Access Development of an efficient protocol for micropropagation of strawberry (Fragaria ananassa Duch.)(CCSHAU, Hisar, 2020-07) Swati Rani; Upendra KumarStrawberry (Fragaria × ananassa Duch.) is the most important fruit worldwide. Micropropagation is mainly for the clonal multiplication.Strawberry cvs. Grenada and Petaluma were micropropagated for rapid shoot and root multiplication. Meristematic part and leaf primordia is source of explants. Foe the sterilization process, the explants were dipped in two to three drops of Tween 20 per 100 ml for 10 minutes with distilled water, 0.5% Bavisitin & Streptocycline for 30 mintues with antimicrobial supplement and 0.1% HgCL2 for 2 minutes gave the maximum aseptic cultures. After the surface sterilization of meristemetic part of plants 3-5 mm long was used as a explant. Multiplication stage results indicate that highest auxiliary buds were observed when MS medium supplemented with 1 mg/l and 1.5 mg/l BAP in Petaluma and Grenada. At rooting stage, it was clear visually that MS medium supplemented with 0.5 mg/l of BAP with 1.5 mg/l of IBA in both the cultivars gave the best results of enhanced number roots with higher length and number shoots with higher length per explants. The best result for root multiplication indicating the use of IBA with ( 1.0, 1.5 mg/l) concentration as compared to other treatments.The highest response for the shoot multiplication was obtained with MS containing 1.5 mg/l and 1.0 mg/l BAP respectively. The present research is very useful in commercializing the new cultivars of strawberry into north Indian conditions which gives higher yield of fruits.ThesisItem Open Access ―Phenotypic and molecular characterization of HD 2967 x Kharchia 65 F5 and backcross generations for salt tolerance in wheat (Triticum aestivum L. em. Thell”(CCSHAU, Hisar, 2019-08) Saini, Pooja; Yashveer, ShikhaSoil salinity is emerging environmental stress that reduces rate of germination, delays emergence and affects almost all stages of growth and development of wheat plants which ultimately leads to reduction in grain yield of crop.The present investigation focused on phenotypic and molecular characterization of selected wheat (Triticum aestivum L. em. Thell)genotypes of F5 and backcross (BC1F4, BC2F3, BC3F1) generations derived from HD 2967 X Kharicha 65 for salinity tolerance on the basis of presence of both Nax loci and their agronomic performance at salt stress level 8 dS/m. A total of 92 plants confirmed the presence of Nax1 locus at a band size of 210 bp consisting of 16 F5, 41 BC1F3, 9 BC2F2 and 27 BC3F1, a total of 53 plants confirmed the presence of Nax2 locus at a band size of 225 bp comprising of 6 F5, 35 BC1F4, 8 BC2F3 and 4 BC3F1.Overall, plants comprising of 6 F5,26 BC1F4, and 4 BC3F1 of HD 2967 x Kharchia 65were found to have both the Nax loci. On the basis of phenotypic and genotypic variations, 56 high yielding plants were selected for genotyping using polymorphic SSR markers. A total of 180 SSR primers were used to study the polymorphism between parents HD 2967 and Kharchia 65, out of which 27 SSR primers showed polymorphism. Polymorphism information content (PIC) value in the studied genotyped ranged from 0.191 (xgwm480) to 0.425 (wmc3) with an average value of 0.339. Cluster tree analysis using NTSYS-pc software, F5 and backcross (BC1F4, BC2F3, BC3F1) generations showed that all the selected plants were inclined toward recurrent parent HD 2967 representing higher similarity with HD 2967. From above analysis, high grain yielding and salt tolerant plants were selected such as plant number 259Q3c, 271Q3c (F5); 202P9b, 303P2b and 202P3b (BC1F4); 241R2c and 243R1a (BC2F3); 25(3)1b and 24(1)1c (BC3F1) and these plants could be further backcrossed with the parent HD 2967 to develop salt tolerant wheat lines.ThesisItem Open Access In silico identification and physical mapping of gene(s) responsible for β-glucan in Bread Wheat (Triticum aestivum. L)(CCSHAU, Hisar, 2020-07) Dhamija, Aryan; Upendra KumarDietary fibers from plant cell wall are an essential component of healthy foods. Higher intake of dietary fiber reduces the risk of diet related chronic disease like type 2 diabetes, obesity and also improves gastrointestinal health. In human diets, cereal fiber is the largest contributor to total dietary fiber consumption. The soluble dietary fiber (1-3) (1-4) mixed linked β-D-glucan from cereal grains is a valuable component of a healthy diet. In the present investigation, the gene responsible for β-glucan in hexploid wheat was identified using both in silico and molecular methods.The in silico analysis confirmed that CslF6 gene in wheat was located on the chromosome 7A. Sequence similarity search was conducted between HvCslF6 and wheat survey sequences displayed a similarity of 96% with chromosome 7A. The structure of this gene had 3 exons and codes for a protein of 945 amino acids. Homology modeling of putative CslF6 protein was described as Probable cellulose synthase A catalytic subunit 8.Different cytogenetic stocks of Chinese spring revealed that CslF6 gene was located on centromeric region of 7AL (FL=0.29). Identified CslF6 in wheat can be utilized to make β-glucan efficient wheat and can be used in biofortification program.ThesisItem Open Access Genome-wide association mapping in barley for terminal heat tolerance and malting quality(CCSHAU, Hisar, 2021-07) Verma, Swati; Yashveer, ShikhaBarley (Hordeum vulgare L.), is one of the major cereal crops and has been cultivated since ancient times in different parts of the world. In particular, heat stress at the post-heading stage causes considerable yield reduction due to the stress. Understanding the genetic variation, changes in physiological processes, and level of genetic diversity existing among genotypes are needed to produce new cultivars not only having a high tolerance to heat stress, but also displaying high yield. So to address this challenge, a panel of 316 diverse barley genotypes (AM2017) were procured. It was evaluated under two conditions timely-sown (TS) and late-sown (LS) conditions in two seasons of 2017-18 and 2018-19 at CCSHAU, Hisar. Ten agro-morphological, four physiological, and five grain malting quality traits were studied. Genetic diversity and population structure were explored in the AM2017 panel after genotyping it with the 50 K iSelect Illumina Barley SNP array. A set of 36,793 SNP markers, covering a genetic distance of 991.82 cM with an average marker density of 37.09 SNPs/cM, was obtained after quality filtration. The gene diversity (GD) and Polymorphic Information Content (PIC) at the genome level were 0.362 and 0.289, respectively. The phenotypic results showed a clear reduction in trait performance under the LS condition. Marker-trait associations (MTAs) were estimated using the compressed mixed linear model. Based on the estimated MTAs and linkage disequilibrium (LD) decay observed in the genome, putative QTL was identified as associated with the traits studied. The most robust QTL observed under both sowing conditions were further studied and validated using the previously reported QTL. Several traits were found to have QTL that was not found to be reported. The combination of analyses using SNPs and changes in physiological traits provided useful information on genomic regions taking part in heat stress tolerance.ThesisItem Open Access Screening of wheat genotypes for multiple fungal resistance using functional markers(CCSHAU, Hisar, 2021-09) Garg, Bharat; Yashveer, ShikhaIn the present study, 58 wheat genotypes were characterized for grain yield, biochemical traits and functional markers. The wheat genotypes were sown in RBD (Randomized Block Design) design in two replications in the field of Wheat and Barley Section, Department of Genetics and Plant Breeding, CCS Haryana Agricultural University, Hisar, Haryana (India) during Rabi, 2019-2020 for the evaluation of ten yield and its component characters viz. plant height, number of tiller per meter, spike length, number of grain per spike, number of spikelet per spike, grain yield per meter, biological yield per meter, harvest index and 1000-grain weight. In this study 58 wheat genotypes were screened for morphological and biochemical traits, showed varied mean range signifying ample amount of variability present in the wheat genotypes Genetic variability of genotypes was assessed by PCV, GCV, heritability and genetic advance over mean. The values of GCV were lower than their corresponding PCV for all the characters studied, indicated the negligible effect of environment on the development of plants. The maximum value of the GCV & PCV was observed for the grain yield per meter (11.87) and number of spikelet per spike (16.65). Highest broad sense heritability was recorded for biological yield per meter (80.92) The positive and significant genotypic correlation for grain yield per meter was found with number of tillers per meter (0.503), spike length (0.217), number of grains per spike (0.332), biological yield per meter (0.864), harvest index (0.616). Thus, these component traits may be used in wheat breeding programme for further improvement in grain yield. For characterization of quality traits, various biochemical attributes were analyzed viz. total phenolic content, total flavonoids, proline content, glycine betaine, phenylalanine ammonia lyase (PAL) and tyrosine ammonia lyase (TAL) at vegetative and maturity stages. Total phenolic content range for vegetative stage is 0.41 to 4.74 mg CE/g and 1.07 to 6.17 mg CE/g in the maturity stage. Total flavonoids range for the vegetative stage, is 0.30 to 0.97 mg QE/g and 1.07 to 9.18 mg QE/g in the maturity stage. Proline content range for vegetative stage is 0.07 to 0.45 mg /g and 0.08 to 0.55 mg /g in maturity stage. Glycine betaine range for vegetative stage is 0.59 to 9.59 mg /g DW and 2.67 to 4.24 mg /g DW in the maturity stage. PAL activity range for vegetative stage is 4.15 to 6.23 μmole/h/g and 4.99 to 6.51 μmole/h/g in the maturity stage. TAL activity range for vegetative stage is 1.45 to 12.43 μmole/h/g and 1.74 to 12.10 μmole/h/g in the maturity stage. Total flavonoids content had high GCV and PCV in biochemical traits. On heritability analysis, Tyrosine ammonia lyase (TAL) had higher heritability. High heritability coupled with high genetic advance was found in characters such as total phenolic content, total flavonoids content, proline content, glycine betaine and TAL. These characters are recommended in selecting genotypes for screening for biotic and abiotic stress. For functional marker analysis, a total of 24 functional markers were used showed amplification. These selected primers are linked with specific fungal disease resistance gene and amplification of specific primer on wheat genotypes gave the idea of presence of that allele.ThesisItem Open Access Molecular approaches for detection and forecasting of wheat yellow rust(CCSHAU, Hisar, 2020-03) Rizwana Rehsawla; Yadav, Neelam R.Yellow or stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is a devastating airborne disease that affects bread wheat in the major wheat growing regions of India. The understanding of the origin, evolution, pathogenicity, avirulence/virulence behaviour of Pst is very important for the development of more effective management strategies to combat the disease spread. To understand inter and intraspecific phylogenetic relationship among Indian Pst pathotypes, multigene sequence analysis was done. Molecular marker study along with sequencing technology was used to collect information, which was more effective than virulence characterization. The molecular diversity analysis among 13 different Pst pathotypes showed two major cluster formations at similarity coefficient of 0.78. Sometimes, all three wheat rust or two rusts in combination occur simultaneously in the field. Under such conditions identification and differentiation of the yellow rust is needed for precise identification and high throughput DNA based detection protocols. To address this problem PCR based markers were developed which can specifically detect and differentiate Pst from two other rust species of Puccinia and other wheat pathogens. DNA-based methods such as conventional PCR have revolutionized plant disease detection; they are not very reliable at asymptomatic stage. Therefore, a simple and reproducible LAMP assay was developed which could detect the pathogen at an early stage i.e. 3rd day of post infection without any visible sign of pathogen attack on the leaf sample using LAMP primers available in public domain. Four novel sets of LAMP primers from ketopantotate reductase gene were also designed for Pst detection which worked successfully. Biosensing by electrochemical and SPR for the detection of yellow rust was also undertaken. Electrochemical based sensing was done using different sequences of Pst specific genes as probes. Linear response over wide DNA concentration range from 10 pg/μl to 115ng/μl was obtained with a high sensitivity, accuracy and reproducibility. The lowest detection limit was observed for microRNA like RNA 1 gene i.e. 10 pg/μl. Electrochemical DNA based biosensing was developed to distinguish between the yellow rust susceptible and resistant wheat genotypes using TaATG8j gene sequence. Linear response over wide DNA concentration range from 1 ng/μl to 50 ng/μl was obtained with detection limit of 4 pg/μl. The SPR biosensor demonstrated high specificity and long shelf life thus promising its application in Pst diagnosis. The developed biosensor exhibited a high sensitivity (0.18°/ (ng/μl)), good linearity, low detection limit (1 ng/μl) and high specificity over a wide concentration range of DNA (1–150 ng/μl).ThesisItem Open Access Characterization of wheat germplasm for grain yield, quality traits and functional markers(CCSHAU, Hisar, 2019-08) Verma, Priyanka; Yashveer, ShikhaIn the present study, 98 wheat genotypes were characterized for grain yield, quality traits and functional markers. The wheat genotypes were sown in RBD (Randomized Block Design) design in two replications in the field of Wheat and Barley Section, Department of Genetics and Plant Breeding, CCS Haryana Agricultural University, Hisar, Haryana (India) during Rabi, 2018-2019 for the evaluation of nine yield and its component characters viz. plant height, number of effective tiller per meter, spike length, number of grain per spike, number of spikelet per spike, 1000-grain weight, grain yield per plot, biological yield per plot. The values of GCV were lower than their corresponding PCV for all the characters studied, indicated the negligible effect of environment on the development of plants. The highest value for GCV and PCV was observed on biological yield per plot (18.09 & 19.18 %) followed by harvest index (15.76 & 16.04 %) and number of grains per spike (14.29 & 14.60 %), respectively. Heritability for broad sense (h2) showed highest for harvest index (96.50 %) followed by number of grains per spike (95.90 %) and biological yield per plot (89%). Genetic advance as 5% of mean was observed highest for biological yield (35.17%) and harvest index (31.89%), indicating a good scope for improvement in grain yield and its component characters. Grain yield per plot was highly and significantly positive correlated with number of effective tillers per plot (0.448), spike length (0.551), number of grains per spike (0.611), number of spikelets per spike (0.588), 1000 grain weight (0.585), biological yield per plot (0.805) and harvest index (0.113). Path analysis revealed that biological yield per plot had the maximum direct positive effect on grain yield per plot (0.600) followed by harvest index (0.573), whereas, all traits contributed indirectly towards grain yield per plot via biological yield per plot except plant height (-0202). The grain yield component traits viz. biological yield per plot, harvest index and number of grains per spike were showing high GCV, PCV, heritability and genetic advance as 5% of mean. Thus, these component traits may be used in wheat breeding programme for further improvement in grain yield. For characterization of quality traits, various biochemical attributes were analyzed viz. protein content, mineral (Fe, Zn), phytates, gluten content, amylose content, amylopectin content, hectolitre weight and sedimentation value. Protein content ranged from 9.4 to 15.2 per cent with mean of 12.2 per cent, iron content from 21.3 to 39.3 mg/kg with mean of 29.3 mg/kg, zinc content from.15.7 to 40.70 mg/kg with mean of 29.60 mg/kg, phytates (μg/g) from 4816.0 to 8757.3 (μg/g) with mean of 7008.5 μg/g, wet gluten content from 8.00 to 37.90 per cent with mean of 24.20 per cent, dry gluten content ranged from 3.00 to 14.11 per cent with mean of 10.96 per cent, total soluble sugar content from 0.86 to 6.16 per cent with mean of 3.05 per cent, starch content from 49.8 to 69.4 per cent with mean of 56.7 per cent, amylose content from 3.02 to 8.45 per cent with mean of 5.48 per cent, amylopectin content from 49.36 to 68.97 per cent with mean of 56.17 per cent, hectolitre weight from 72.0 to 87.30 kg/hl with mean of 80.10 kg/hl, sedimentation value from 19 to 41 ml with mean of 31.2 ml. For functional marker analysis, a total of 6 functional markers were used showed amplificationThesisItem Open Access ISSR Markers Based Genetic Diversity Analysis in Kiwifruit (Actinidia deliciosa (A.Chev.) CF Liang et AR Ferguson) Genotypes(CCSHAU, Hisar, 2019-09) Jhilta, Agrim; Kharb, PushpaKiwifruit (Actinidia deliciosa (A. Chev.) CF Liang et AR Ferguson) (2n=6x=174) also known as Chinese goose berry and the horticultural wonder of New Zealand, is a commercially important crop and a very good source of Vitamin C. The present study was undertaken for assessing genetic diversity in Kiwifruit genotypes using ISSR primers and for identification of any probable ISSR marker linked to sex in Kiwifruit genotypes. A total of seven genotypes of Kiwifruit were used that included two male (Tomuri and Allison (M)) genotypes and five female genotypes (Allison, Abbott, Bruno, Hayward, Monty). Genomic DNA was isolated from young leaves of Kiwifruit genotypes. For molecular analysis 104 ISSR primers were screened, out of which 24 ISSR primers amplified and were polymorphic. These were employed for genetic diversity analysis and identification of genotype. Among these polymorphic ISSRs, eight ISSRs were found to identify selected genotypes on the basis of unique amplicons. IS7, IS11, IS93, IS88, IS114, IS107, IS101 and IS79 could identify genotypes . One SSR marker, A 003 reported earlier was also validated. It could identify Allison (M). Similarity matrices of seven kiwifruit genotypes revealed that similarity value among different genotypes ranged from 0.5298 to 0.7094. A maximum similarity value of 0.7094 was observed between Allison (M) and Tomuri whereas Abott and Allison (F) were found to be most diverse. Molecular analysis showed that Allison (M) and Tomuri were much similar. Abbott was found most diverse when compared to other kiwifruit genotypes. The results of present study can be utilized for sex and genotype identification at seedling stage and in kiwifruit improvement programme.