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ThesisItem Restricted Development and validation of kaspmarkers for heat stress tolerance QTLs in wheat (Triticum aestivum L.)(Punjab Agricultural University, Ludhiana, 2022) Ranjeet Singh; Dhaliwal, InderpreetA panel of 24 wheat genotypes, comprising 19 heat tolerant, 3 heat susceptible and 2 parental lines were evaluated under timely and late sown conditions during rabi 2020 in a Randomized block design with an aim to develop and validate a KASP assay for heat stress tolerance. Eleven major reproductive traits, including grain yield, plant height and days to maturity were evaluated. In general, there was a reduction in grain yield under late sown compared to timely sown conditions. This trait was highly variable among the 24 genotypes as some of them (KSG1188, KSG089, KSG849, KSG1189 and KSG1195) exhibited very low reduction in yield, indicating a tolerance to heat stress for maintaining the desired yield levels. Similar behavior was observed for other plant traits also, viz, days to heading, days to maturity, mean grain weight per main spike, spikelet fertility and tiller number per meter square. There were also some genotypes that displayed increased height and biomass under late sown conditions, which is rare for a heat stressed crop. The stability in performance of the genotypes may be attributed to heat stress tolerance under late sown conditions. Since the QTL for stress tolerance were already described in an earlier study, the identified GBS-SNPs were converted to 4 KASP primers following standard procedures to run the KASP assay on this panel. Of these four primers, 2 were successful in allele calling, thereby giving an efficiency of 50%. Hence, converting these identified GBS-SNPs into KASP assays can successfully be used to differentiate the genotypes or populations for heat stress related QTL.ThesisItem Embargo Phenotypic and molecular characterization of rice (Oryza sativa (L.)) genotypes for grain zinc content(Punjab Agricultural University, 2022) Phyo, Su Hlaing; Mangat, G. S.Biofortification is one of the most sustainable strategies to alleviate micronutrients deficiencies for people who mainly consume rice and have limited access to diversified food. The present study aimed at evaluating a set of 160 advance breeding lines along with check varieties for various yield attributes, quality traits and grain zinc content at Ludhiana and Rauni. These were evaluated for days to 50 % flowering, plant height, ear bearing tillers per square metre, spikelets sterility, number of spikelets per panicle, thousand grain weight and grain yield, milling quality and grain zinc content. Significant genotypic variation was present for various traits at both locations. Most of the genotypes were early to medium in maturity with semi dwarf plant height and good tillering ability. Significant variation for head rice recovery was present and it ranged from (41.7 to 67.6%). The zinc content in brown rice ranged from 17.1 to 35 ppm and in polished rice from 12.3 to 29.7 ppm. The genotypes PAU 8182-205-2, PAU 8182-205-1, PAU 8182-108-1, PAU 8182-116-1, PAU 8182-150-1, PAU 8182-182-1, PAU 8182-112-5, PAU 8182-171-4, PAU 8182-175-1 and PAU 8182-215-1 had high grain zinc content of more than 24 ppm in polished rice. Grain yield was negatively correlated (-0.208** and -0.185*) with grain zinc content in brown rice and polished rice. Molecular characterization of advance breeding lines with known markers for high zinc contents did not reveal much variation as most of the primers showed monomorphic banding pattern. Zinc rich elite genotypes with desirable ancillary traits may serve as potential donors for biofortification breeding programme.ThesisItem Restricted Senescence, nutrient remobilization and yield in wheat germplasm introgressed with Gpc-B1 gene(Punjab Agricultural University, Ludhiana, 2022) Jafar Tanin, Mohammad; Sharma, AchlaThe present breeding efforts are required to evaluate a set of germplasm with the Gpc-B1 gene in their genetic background. However, Gpc-B1 is the only recognized and cloned gene which explains 66% of the phenotypic variation for GPC. Therefore, it is essential to enrich our knowledge regarding its role in protein accumulation in wheat grain. On the other hand, the Gpc-B1 has a negative relationship with grain yield. This has made it difficult to create an elite wheat line that has both high GPC and a high yield. The accelerated senescence due to the presence of this gene shortens the grain filling period. We phenotyped the NILs for different phonological and yield-related traits, including number of days to flowering (DTF), plant height (PH), number of spikelets per spike (SPS), 1000-grain weight (TGW), yield, number of days to maturity (DTM), and grain protein content (GPC). An alpha-lattice design was used to do an analysis of variance, which showed that there were significant differences between the genotypes for all of the traits. The Gpc-B1 gene accelerated the flowering time and maturity time by about one day and two days, respectively, as compared to genotypes having no Gpc-B1 gene in their genetic background. Simultaneously, the same gene made the plant shorter by an average of 4.41 cm. The functional Gpc-B1 gene in the genetic background of the NILs increased the protein accumulation in wheat grain through an accelerated senescence and more efficient nitrogen transportation process, which was initiated earlier. But at the same time, the improvement in GPC could result in a grain yield penalty due to a reduction in grain size. A better understanding of how nutrients or chemicals change the function of wheat plants could help breeding programmes in the future. The NILs were evaluated under KNO3 and salicylic acid treatment and control (no KNO3 and salicylic acid application) conditions with the standard fertilizer application rate recommended by PAU for one year (2020–21). A combined analysis of variance using an alpha lattice design was conducted, which indicated significant variation among the studied traits, including DTF, PH, SPS, GPS, DTM, HI, GPC, Yield, and TGW. The potassium nitrate application significantly affected the PH, GPS, DTM, TGW, yield, HI, chlorophyll content in flag leaf at anthesis state and 30 DAA, and other traits. Salicylic acid application significantly affected the number of grains per spike (GPS), 1000-grain weight (TGW), yield, and number of days to maturity (DTM) as compared to the control experiment. This delay had no effect on GPC in NILs, but it did increase yield and traits related to yield, like TGW and GPS. A pooled analysis of variance using an alpha lattice design was conducted, which showed significant variation for all the traits under study. High GPC variation in lines with the same genetic background (PBW550) led to a lot of questions. To find the reasons for this variation, we isolated the high quality RNA of two high GPC NILs and two low GPC NILs. Differential gene expression analysis was carried out on each NIL as a control. A total of 1418 differentially expressed genes (DEGs) were discovered for the selected NIL across all three stages, out of which 1117 genes were upregulated while 301 genes were downregulated. This suggests their probable role in seed development.ThesisItem Restricted Identification of SSR markers associated with Plant Architectural Traits in cowpea (Vigna unguiculata L. walp)(Punjab Agricultural University, 2022) Jain, Mohit; Devinder Pal SinghThe objective of the present investigation was to estimate genetic variation and type of gene action controlling morpho-agronomic traits of two cowpea crosses viz., C-88 x TNFC 6926 and CL400 x C-74. The data for six populations viz., Parental lines their FI, F2, BC and BC were collected for evaluation. The observations were recorded for different morphological attributes viz; plant height (cm), vine length (cm), number of leaves, leaf length (cm), leaf width (cm), green fodder yield (kg/plot), dry matter yield (kg/plot), crude protein (%), acid detergent fibre (%), neutral detergent fibre (%) and in vitro dry matter digestibility (%).Simple- additive model was tested to check the adequacy and presence of epistatic interactions. The scaling test shows significant results for most of the traits for both crosses which indicated the presence of epistatic interactions. When the simple additive - dominance model was inadequate, the data were subjected to the six parameter model which included digenic epistatic effects. The results indicated that the mean effect is highly significant and the traits were quantitatively inherited. Additive and dominant gene effects are highly significant for most of the traits under investigation. The high magnitude of additive x additive gene effect for green fodder yield suggests the pedigree method best suitable breeding programme for development of cultivars with better fodder quality traits. ldentification of SSR primers associated with plant type was conducted on F: population of Cross I (C-88 x TNFC 6926). 151 SSRs were employed for Bulk Segregant Analysis (BSA), out of which 15 SSRs showed polymorphism. These SSR primers can be utilised in future for the selection of semi-erect plant types from large Fa population. Semi-erect plant types can be grown along with other non-legume crops for getting nutritious fodder.ThesisItem Restricted Morphological characterization and assessment of genetic diversity in interspecific derivatives of chickpea (Cicer arietinum L.)(Punjab Agricultural University, 2022) Vadithya, Amool Singh; Inderjit SinghIn the present investigation, a set of 90 interspecific derivatives, derived from four different crosses between cultivated chickpea (PBG5 and BGD72) and wild species (Cicer reticulatum ILWC229 and C. echinospermum ILWC229) were evaluated along with their parents and two standard checks (PBG7, PBG8) to determine the variability, association, divergence and resistance to Ascochyta Blight and Botrytis Grey Mould diseases during 2020-21 and 2021-22 in the experimental area of pulses section, PAU, Ludhiana. The genotypes were planted in αlattice design with two replications. ANOVA revealed significant differences among the derivatives for all the traits studied. Phenotypic and genotypic coefficients of variation were high for the number of pods per plant followed by primary and secondary branches per plant and 100-seed weight. High heritability estimates were observed for plant height followed by days to flowering, harvest index, days to maturity, number of pods per plant and biological yield per plot. Seed yield per plot had a positive and highly significant correlation with biological yield per plot, plant height, harvest index, 100-seed weight and the number of pods per plant. Path analysis revealed highest positive direct effect of biological yield per plot on seed yield per plot, followed by harvest index, secondary branches per plant, days to 50% flowering, 100-seed weight and days to maturity. Based on genetic diversity analysis, 90 derivatives were grouped into 5 clusters. The maximum inter-cluster distance was observed between clusters II and V. Cluster II was good with respect to seed yield and biological yield per plot, 100-seed weight and days to 50% flowering and maturity. A total of 33 derivatives were found to be resistant to AB disease and 12 were found to be moderately resistant to BGM disease. The parental lines were screened for polymorphism using 450 SSR markers. The introgression from C. reticulatum and C. echinospermum was confirmed by graphical genotyping. Resistance to AB was confirmed by markers viz. TR19, TA194, TA96, TA106 and H1I16 and resistance to BGM by TA194, TA140 and TA118. In addition, TA113 and TA18 were related to higher seed weight and tallness, respectively. Therefore, these markers can be used with sufficient confidence in marker-assisted selection to develop superior lines for resistance to AB and BGM along with higher seed weight and plant height.ThesisItem Restricted Identification and characterization of QTLs governing key nitrogen fixation and nutrient traits in chickpea (C. arietinum L.)(Punjab Agricultural University, Ludhiana, 2022) Sethi, Sorabh; Sarvjeet SinghThe present study was conducted with an objective to identify and characterize the genetic factors underlying two broadly categorized traits viz., nutrient availability and nitrogen fixation related traits in chickpea and their relevance with economic yield in chickpea. The analysis of variance (ANOVA) revealed significant variation for all the traits under study among all genotypes across the years and environments (location) and also in pooled data. Measurement of skewness and kurtosis indicated presence of duplicate and complementary gene interaction for different traits. Correlation analysis indicated positive and significant correlation of seed yield with pods per plant. Similar association was observed between days to 50 % flowering and days to maturity. Apart from agronomic traits, no significant correlation was observed between nutrient and nitrogen fixation related traits. However, significant and positive correlation was observed among Fe and Zn content traits and also among different nitrogen fixation traits, but the pattern was not consistent for all the years and locations. For genotypic analysis, genotypic data was generated using genotyping by sequencing (GBS-ddRAD) approach. Total of 25,233 filtered SNPs were obtained based on 2X genomic coverage but post filtering, 520 SNPs were eventually used for generation of linkage mapping and bi-parental QTL mapping using QTL ICi mapping software. Total 14 QTLs have been identified, out of which four QTLs have been identified for nutrient traits i.e., three for grain Fe content while one QTL for Zn content. Six QTLs were identified for nodule number trait while for leghemoglobin content, total of four QTLs identified. In our study, no statistically significant QTL were identified for chlorophyll content and nodule dry weight. Promising lines for grain iron, zinc content and nitrogen fixation traits were identified which can be used as donor parents in chickpea breeding programme and developed markers can be used in marker assisted selection (MAS) for identifying desirable genotypes having allele of interest.ThesisItem Restricted Characterization of introgressed Gossypium hirsutum lines for cotton leaf curl disease resistance(Punjab Agricultural University, Ludhiana, 2022) Gurcharn Singh; Paramjit SinghThe plant material consisted of BC2F4 progenies obtained from G. hirsutum cv. F 1861/G. armourianum acc. PAU 1/G. hirsutum cv. LH 1556 cross. This material was evaluated for CLCuD response and several other economic traits. For inheritance of CLCuD resistance and identification of SSR markers linked to CLCuD resistance, a BC5F2 population derived from the cross: G. hirsutum cv. F1861/ G. armourianum acc. PAU 1/*5G. hirsutum cv. F 1861 was used. A set of 121 backcross progenies including F 1861, LH 1556 and F 2228 were planted during kharif 2020 in a simple lattice design (11 x 11) with two replications. A set of 85 cotton specific SSR markers were used for documenting polymorphism between the parents and genotyping the mapping population. Six progenies were observed to be completely free from the disease, 76 progenies were segregating for CLCuD reaction, whereas, 36 backcross progenies were found to be completely susceptible to CLCuD. A simple genetic control for CLCuD resistance (monogenic dominant) has been observed in this material. Significant differences among the cotton genotypes were observed for seed cotton yield, upper half mean length, uniformity index, tenacity and micronaire indicating substantial variation for these characters in the experimental plant material. Mean seed cotton yield plant-1 ranged from 46 g (Progeny 53) through 129.5 g (progeny 298). None of the 118 backcross progenies could significantly out yield standard check F 2228 (126 g), though one progeny 298 produced numerically higher (129.5 g) seed cotton yield than that of the check. Mean fibre strength ranged from 24.9 g/tex (Progeny 567) through 32 g/tex (progeny 113). Sixty four progenies significantly outperformed the check F 2228 (26.65 g/tex). Mean micronaire value ranged from 3.85 (Progeny 108) through 5.4 (progeny 412). Most of the backcross progenies significantly outperformed the standard check F 2228 (5.3) for this trait. One SSR marker (NAU 3903) was found to be polymorphic between the resistant and susceptible F2 bulks.ThesisItem Embargo Response of Null Kunitz Trypsin Inhibitor Soybean (Glycine Max L. Merrill) Genotypes to Different Environments(Punjab Agricultural University, Ludhiana, 2022) Nge, Su Yadanar; Dhillon, Satwinder KaurThe present study entitled “Response of Null Kunitz Trypsin Inhibitor Soybean (Glycine max (L.) Merrill.) Genotypes To Different Environments” consisting of 27 genotypes involving 21 null-Kunitz trypsin inhibitor lines, and four check varieties, two parents i.e SL 525 and NRC 101 was conducted in year 2021 in four different environments i.e two locations over two sowing dates, E1 (June sowing at Ludhiana), E2 (July sowing at Ludhiana), E3 (June sowing at Gurdaspur), E4 (July sowing at Gurdaspur). Analysis of variance showed significant differences among genotypes, environments and their interactions. Genotype x Environments interaction and stability analysis revealed significant differences among the genotypes for all eleven traits studied except seeds per pod over environments, representing the presence of considerable amount of genetic variability in experimental materials. The mean squares due to environment (linear) were significant for all traits. Non-linear component of G x E interaction was significant for all traits. Genotype x environments (linear) were significant for plant height, number of pods per plant, 100 seed weight, seed yield and trypsin inhibitor activity. A wide range of mean values was recorded for the traits viz. 50% flowering (50-65 days), day to maturity (110-126 days), plant height (56.5-120.43cm), number of branches per plant (2-4), number of pods per plant (26-37), number of seeds per pod (2.26-2.65), hundred seed weight (7.6-11.32g), seed yield per plant (6.73-11.33g), oil content (22.3-19.2%), protein content (39.4-41%) and trypsin inhibitor content (9.29-66.86%) in all environments. The genotypes KTI-13, KTI-41, KTI- 52, KTI-59, KTI-64 (absence of KTI gene) and KTI-30, KTI- 31, KTI- 123, KTI- 115, KTI-156, KTI-157 (presence of KTI gene) exhibited stability for superior performance w.r.t. seed yield and its important components in all the environments. Amongst a total of 21 genotypes only 11 showed absence of KTI and 10 genotypes recorded presence of KTI. Phenotyping was done by measuring trypsin inhibitor activity (TIA). Consequently, the genotypes of Null KTI gene recorded reduced levels of TIA.ThesisItem Embargo Evaluation of germplasm and expression analysis of TFL1 gene for stem growth habit in urdbean(Punjab Agricultural University, Ludhiana, 2022) Baljeet Kaur; Ashok KumarThe present study consisted of one hundred genotypes of urdbean collected from different sources to study the stem growth habit, flowering duration and expression analysis of genes responsible for stem growth habit during different developmental stages. The experiment was conducted during spring season in 2021 at two locations (PAU, Ludhiana and PAU, RRS, Gurdaspur) in simple lattice design (10x10) using two replications. The analysis of variance showed that all the characters studied exhibit significant variation at both locations. GxE interactions were also found significant for all the characters. At Ludhiana, the characters like Plant height at first pod maturity (H2), Plant height at 90% pod maturity (H3), Number of pods per plant, Biological yield per plant, DDh1-3, DDh2_3 and DDd4_5, at Gurdaspur, Plant height at 90% pod maturity (H3), Number of pods per plant, Biological yield per plant, DDh2_3 and DD4_5 and pooled data analysis over the locations showed that Plant height at 90% pod maturity, Number of pods per plant, Biological yield per plant, yield per plant, DDh2_3 and DD4_5 possessed high PCV, GCV, heritability and GAM. The genotypic variability for all these characters are probably due to predominance of additive gene action and can be improved through phenotypic selection. The variability for these characters present in the germplasm could be exploited for developing early maturing, lodging resistant and high yielding varieties of urdbean. The expression analysis showed that TFL1 gene at three leaf stage in both the accessions i.e. IC569464 (determinate) and IC530625 (indeterminate) are similar. The expression was observed highest in flowering stage, first pod maturity and 90% pod maturity in indeterminate growth habit over determinate of plant. Specific primers were designed to check the expression of candidate gene in determinate growth habit and indeterminate growth habit at different growing stages of the plant. Hence, present study helped us to understand the molecular mechanisms of genes underlying in determinate and indeterminate growth habit of the plant in urdbean.
