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ThesisItem Restricted Evaluation of promising lines for yield and quality traits along with leaf curl virus disease resistance in pickled chilli (Capsicum annuum L.)(Punjab Agricultural University, 2024) Ravinder Singh; Dr S.K. JindalThe present study was conducted to evaluate seventy-five pickled chilli genotypes for various yield and quality traits at Vegetable Research Farm, Punjab Agricultural University, Ludhiana during 2021-23. The research findings depicts that all the genotypes differ in the expression of different traits. Like, genotype PCP-732-2 exhibited the highest plant height (72.1 cm) but was not superior in other traits. All the characters evaluated, possessed high heritability along with high genetic advance as a percent of the mean except for number of fruits per plant. Fruit yield per plant exhibited significant and positive correlation with fruit length, fruit width, pericarp thickness, number of fruits per plant and average fruit weight at both the phenotypic and genotypic levels. Path analysis had provided insights that number of primary branches, fruit length, pericarp thickness, number of fruits per plant, average fruit weight, capsaicin in green fruits, oleoresin content and colouring matter have direct and positive effect on fruit yield per plant at both phenotypic and genotypic levels. Therefore, emphasizing these Principal component analysis indicated that most of the variation among the genotypes was contributed by the first seven principal components (> 70%). Among the variables, plant height emerged as a key contributor followed by the number of primary branches, fruit length and fruit width. Cluster analysis concluded that seventy-five genotypes were classified into four clusters. Cluster I entailed maximum number of genotypes (37), followed by Cluster II (30), III (7) and IV (1). The maximum intra-class distance was found in cluster I followed by cluster II and cluster III. The inter-class distance was found highest between cluster I and cluster IV followed by cluster II-cluster III. The larger the inter-class distance, the more distinct and well-separated the clusters from each other. Natural and artificial screening against chilli leaf curl virus disease reported that genotypes specifically, FLP-649, PCP-7001, PCP-701-1 and PCP-754 were found to be resistant while, artificial screening against rootknot nematodes reported that genotype PUP-607 and PCP-701-1 exhibited resistant reaction. Hence, genotype PCP-701-1 possessed resistance against both diseases.ThesisItem Restricted Morphological, biochemical and molecular characterization of available germplasm and induced mutants in garlic (Allium sativum L.)(Punjab Agricultural University, 2023) Karanveer Kaur; Sharma, MadhuGarlic (Allium sativum L.) a bulbous crop propagated through vegetative means, has high therapeutic value due to abundance in sulphur containing compounds. In spite of clonal propagation, garlic has shown tremendous genetic variation. Present study was planned with the vision to explore magnitude of variation at genetic level among 139 garlic clones. Significant variation at morphological, biochemical and molecular level was witnessed among the garlic clones. Mahalanobis D2 categorized them into 8 clusters and PCA compressed 15 vectors to 7 components with 92.44% variation. Genotyping analysis represented that the PIC value ranged from 0.041 - 0.888, with average value of 0.583. Jaccard’s dissimilarity coefficient constructed dendrogram into two major clusters, where garlic clones belonging to Allium ampeloprasum formed cluster-I and rest Allium sativum subsp sativum and Allium sativum subsp ophioscorodon formed cluster-II. Extensive variation in biochemical attributes among various genotypes exhibited huge variation, where, dry matter ranged from 26.68 to 53.96%, pyruvic content 119.99 to 443.19mg/100g, allicin content 0.12 to 0.42%, total soluble solid content (TSS) 23.70 to 49.63ºB, total phenol content 0.32 to 1.39mg/100g, ortho-dihydroxy phenol 0.02 to 0.18mg/100g and Flavonol content 0.04 to 0.30 mg/100g. Screening of germplasm against stemphylium blight under natural conditions categorized it into- moderately resistant (28), moderately susceptible (72), susceptible (33) and highly susceptible (6), whereas, no genotype was found to be immune and resistant. Assessment of available germplasm has its own constraints, as it is limited to studying the existing variations and lacks the ability to generate novel genetic alterations. Due to sexual sterility and inability to exchange genetic material, new genetic modifications can be effectively induced through mutation in garlic. Hitherto, two garlic clones exposed to different mutagens viz., EMS, sodium azide and gamma irradiation at various doses and durations gave morphological mutants in MV2 generation, to be utilized in further crop improvement programmes. To overcome the challenge posed by its sterile behaviour, various growth regulators were tried for induction of flowering. Though, these treatments altered the morphology of garlic genotypes, affected aspects like germination as well as early and high scape length but were unable to trigger the process of flowering in garlic. However, the variability available in germplasm provided substantial opportunity to develop new clones with table and processing properties.ThesisItem Embargo Inheritance of fertility restoration in Indian onion (Allium cepa L.)(Punjab Agricultural University, 2024) Kavita; Dhatt, Ajmer SinghOnion is a cross-pollinated crop and is suitable for heterosis breeding to develop elite F1 hybrids. The small size of onion flowers hinders the use of manual emasculation and pollination techniques but could be alleviated by adopting the mechanism of cytoplasmic male sterility. The development of A (Smsms) and B (Nmsms) lines is time consuming and laborious process through conventional approaches but can be accelerated through marker-assisted selection. The existing molecular markers can accurately predict cytotypes, but have been found inadequate in predicting the Ms locus in Indian populations. This indicated the possibility of another fertility restoration locus (Ms) in Indian onion populations. To explore this, the study titled "Inheritance of fertility restoration in Indian onion (Allium cepa L.) was conducted for two consecutive years- Rabi 2021 and Rabi 2022. The parental lines namely, male sterile (D-121-A), maintainer (D-121-B) and restorer (R-230) lines were characterized through morphological, microscopic, and molecular analyses. The visual assessment indicated that male sterile plants exhibited shrunken or distorted anthers in A-line, while male fertile plants displayed plumpy anthers in B and R-lines. Subsequent microscopic analysis using acetocarmine validated these observations, as the sterile plants showed no staining, whereas the anthers of male fertile plants exhibited a distinct red coloration. To conduct segregation analysis, a segregating F2 progeny of the cross-involving CMS A-line (D-121-A) and the R-line (R-230) was used. Phenotyping and chi-square tests conducted on the F2 population indicated a deviation from the expected Mendelian ratio of male fertile to male sterile (3:1). The observed ratio demonstrated a good fit in a 9:7 ratio, suggesting the presence of two dominant complementary genes regulating the fertility restoration locus (Ms) in Indian population. Cytotype markers based on gel electrophoresis (accD and MKFR) and the nuclear-specific marker (AcPMS1) demonstrated accuracy in predicting the N/S cytotypes and Ms locus in the parental lines. However, when genotyping was done in F2 population with AcPMS1, it was observed that out of 160 individuals, 22 plants did not accurately predict the Ms locus. The genotypes of these 22 individuals did not match with their phenotypic appearance. This discrepancy suggested the potential existence of a recombination event between the marker and the QTL (Ms locus). Furthermore, gel-based markers were found to be time-consuming and had low throughput. Consequently, a high-throughput KASP assay was devised to identify cytotypes and the fertility restoration locus (Ms). The KASP based cytotype markers (OC_17 and OC_19) and SNP based nuclear marker (OMs_1 and OMs_2) were utilized to distinguished the cytotypes and Ms locus in parental lines, respectively. OC_17 and OC_19 accurately predicted the cytotypes of A, B and R parental lines and showed complete resemblance with gel-based markers. However, the SNP-based markers used in the F2 population showcased varying results. OMs_1 displayed clear and distinct cluster formation, while OMs_2 showed distorted polymorphism in the F2 population. The accuracy percentages were 84.82% for OMs_1 in predicting the nuclear Ms locus. The developed KASP based marker (OMs_1) has comparable marker efficiency with the gel-based marker (AcPMS1). As Pran a triploid Allium species is considered as a source of S cytoplasm based on restriction assays in this investigation both gel and KASP-based cytotype markers identified 'Pran', along with A and R-lines, as S-cytoplasm. This provides further support for the idea that the sterile cytoplasm in Indian onion populations possibly introduced from the "Pran". This study being the first report regarding fertility restoration control in Indian onion further emphasizes on mapping genomic regions governing fertility restoration for generating tightly linked markers to enhance marker-assisted breeding programs in Indian onions.ThesisItem Restricted Development of monoecious lines in muskmelon (Cucumis melo L.) through marker assisted selection(Punjab Agricultural University, 2024) Nidhi; Sharma, Sat PalIn heterosis breeding, monoecious sex expression offers significant advantages over andromonoecy in muskmelon, as it eliminates emasculation and produces fruits with desirable traits, such as earliness, high TSS, thick flesh, small scar size, and large fruit size. Thus, development of stable monoecious lines possessing desirable horticultural traits is critical for hybrid breeding in this crop. This investigation was aimed to identify molecular markers linked to monoecious trait and to transfer this trait to a popular melon cultivar through marker assisted selection. For this purpose, stability of monoecism was assessed under varying environments and identified 18 stable monoecious using the andromonoecious index. A set of identified lines was used to develop KASP markers for the sex-determining gene, CmACS-7, which revealed two mutations in exons 1 and 2, with one SNP at 170th, followed by insertion of ‘GA’ at 222nd nucleotide position. Upon validation, KASP-CT-2 accurately separated homozygous andromonoecious and monoecious parents and was also successfully validated on F2 population segregating for monoecious trait. Subsequently, marker-assisted backcross breeding was carried out to transfer the dominant gene 'A' controlling the monoecious trait into the Punjab Sunheri, a popular muskmelon cultivar. Heterozygous plants in BC1F1 and BC2F1 and homozygous plants in BC2F2 with high recurrent parent genome recovery and monoecious trait were advanced using the cleaved amplified polymorphic sequence (CAPS) marker. In the BC2F2 population, homozygous monoecious plants were identified using the CAPS marker. These plants were self pollinated to generate a BC2F3 progenies, of which 11 out of 13 lines had phenotypic similarities with the recurrent parents. These 11 lines were self pollinated to generate 34 BC2F4 progenies. Eight BC2F4 lines, viz. Mono-6R2, Mono-IOB1, Mono-2OB3, Mono-7R3, Mono-2OV2, Mono-1OB2 were selected based on TSS, TSS/TA ratio, β-carotene, low titratable acidity, pH, flesh thickness, fruit shape index, and yield. The information and developed associated molecular marker will be useful in developing new stable monoecious lines. Furthermore, the generated monoecious lines can potentially be utilized as a female parent for F1 hybrid development in muskmelon.ThesisItem Restricted Screening of Abelmoschus species for resistance against yellow vein mosaic disease and root-knot nematode for the development of interspecific F1 hybrids(Punjab Agricultural University, 2023) Thakur, Yukta; Pathak, MamtaThe current study was conducted for evaluation of okra wild species against resistance/tolerance to YVMV disease, root knot nematode, and development of interspecific F1 hybrids with resistant donor parents. Further, parents and their interspecific F1 hybrids were morphologically and biochemically characterized. Sixty-five accessions of Abelmoschus moschatus and seven cultivars of Abelmoschus esculentus were subjected to artificial screening for yellow vein mosaic disease (YVMD) and root-knot nematode resistance. Sequencing of initial source plant (Pusa Sawani) exhibiting YVMD symptoms showed the presence of begomovirus okra enation leaf curl virus (OELCuV). The artificial screening identified twelve highly resistant A. moschatus accessions for YVMV disease, which were later found to be symptomless carriers of the virus upon validation with begomovirus-specific primers (DengF/DengR and OY2395F/OY680R). Five A. moschatus accessions exhibited moderate resistance to root-knot nematodes. Interspecific hybridization between YVMV-susceptible okra cultivars and YVMV-resistant A. moschatus accessions resulted in the development of three interspecific F1 hybrids (Punjab Padmini x EC316073, Pusa Sawani x EC316073 and Punjab Padmini x EC329390). Their hybridity was confirmed using SSR primer Okra037, and these hybrids remained free from YVMD symptom after 90 days of artificial screening, indicating dominant gene control of resistance. Seeds obtained from four interspecific hybrids between root-knot nematode-susceptible and root-knot nematode-resistant parents failed to germinate, indicating lack of compatibility between the parents. Both YVMD and root-knot nematode resistant genotypes depicted elevated levels of phenylalanine ammonia-lyase (PAL), tyrosine ammonia lyase (TAL), peroxidase (POD), and phenolic compounds as compared to susceptible ones and significant negative correlation with disease severity (%) and number of galls per root. This suggests that higher levels of these parameters are associated with resistance to both pests.ThesisItem Embargo Development of begomovirus resistant hybrid(s) using KASP markers in pumpkin (Cucurbita moschata Duchesne)(Punjab Agricultural University, 2023) Sood, Anushka; Sharma, MadhuThe present investigation entitled, “Development of begomovirus resistant hybrid(s) using KASP markers in pumpkin (Cucurbita moschata Duchesne)” was conducted in the Department of Vegetable Science, Punjab Agricultural University, Ludhiana with the objectives to screen pumpkin genotypes using Cmo4018182 KASP marker against mixed infection of squash leaf curl China virus and tomato leaf curl New Delhi virus (MI-Sq/To) and to develop begomovirus resistant hybrids for year-round cultivation of pumpkin. Forty genotypes of pumpkin were screened in spring season 2022, genotypically with Cmo4018182 KASP marker for resistance against MI-Sq/To. Based on KASP marker analysis, nineteen genotypes were selected and randomly hybridized to develop twenty-nine F1 hybrids. In rainy season 2022, the forty genotypes and twenty-nine F1 hybrids were subjected to whitefly mediated artificial mass screening for resistance against MI-Sq/To at nursery stage and 75 days after sowing (DAS). Among forty genotypes, four genotypes were observed as resistant, thirteen as moderately resistant, three as moderately susceptible and twenty genotypes as susceptible at nursery stage. Most of the genotypes showed same disease reaction in nursery stage and 75 DAS except P-37 which later on shifted its reaction from moderately resistant to moderately susceptible. Among 29 hybrids, two hybrids were found resistant, sixteen were moderately resistant, two were moderately susceptible while nine were susceptible at nursery stage. Likewise parents, hybrids retained their disease reaction from nursery to 75 DAS except H9. However, marker trait association of Cmo4018182 KASP marker against resistance to MI-Sq/To was found to be 75% in the diverse panel of 40 genotypes while it was 86.20% in case of hybrids, suggesting further exploration of the identified genomic region for the development of gene-linked markers. In addition to assessing begomovirus resistance, the hybrids underwent evaluation concerning morphological, yield and yield-attributing traits. Significant differences among hybrids for all horticultural traits were observed during both rainy and spring seasons as shown by the analysis of variance. In terms of individual performance, the majority of hybrids displayed substantial improvements in favorable directions for all traits compared to their parents. High genetic variability among genotypes for majority of traits offers an opportunity to develop heterotic hybrids because of their high heritability and genetic advance. Traits such as days to 1st male flower, days to 1st female flower and days to 50% female flower exhibited low genetic variation indicating that hybridization of divergent genotypes could be beneficial in generating diversity in these traits. The significant positive correlation of traits viz, average fruit weight, vine length, number of fruits per plant, fruit diameter, peduncle length, petiole length and flesh thickness with fruit yield indicates the potential for enhancing yield by focusing on these specific traits. Among the 29 hybrids, H8, H18, H20, H24 and H27 were found as the most optimal hybrids based on their resistance to begomovirus, earliness and yield. However, conducting multi-location trials is imperative to affirm the commercial potential of these hybrids. Conclusively, this study showcased the pragmatic value of employing marker assisted breeding in the introgression of begomovirus resistance and expedited the development of begomovirus-resistant hybrids in a shorter duration, conserving both labour and resources.ThesisItem Embargo Studies on growth and yield of garlic (Allium sativum L.) as affected by grading and thermotherapy(Punjab Agricultural University, 2023) Kiranpreet Kaur; Sharma, MadhuThe current study entitled "“Studies on growth and yield of garlic (Allium sativum L.) as affected by grading and thermotherapy”, was conducted with the objectives to generate standard sized virus free propagating material at the Department of Vegetable Science, Punjab Agricultural University, Ludhiana during rabi 2019-21. To standardize the garlic seed cloves for maximum production, four grades viz. grade-A (>12 mm), grade-B (9-12 mm), grade-C (7-9 mm), grade-D (<7 mm) were evaluated against farmer‟s practice of ungraded cloves as control. Grade-A and grade-B outperformed all the grades in terms of growth and yield parameters while, grade-D was the poorest. For production of virus free propagating material, cloves of garlic cv. PG-18 and PG-62 were exposed to 5 hot air treatments (55-60°C) and 7 hot water treatments (55-65°C) for different time spans. Per se performances showed that the first generation (G1-19) of thermotherapy treated cloves performed poorly across treatments in comparison to control for growth and yield parameters in both the garlic cultivars under the influence of high temperature. DAS- ELISA test confirmed 100% virus elimination in four treatments viz. T2(HA2) (55°C for 2 hours), T5(HA5) (60°C for 30 minutes), T6HW1 (55°C for 10 minutes) and T7HW2 (55°C for 20 minutes). The second generation (G2-19) of these four virus free treatments showed normalcy for growth and yield parameters, and T2(HA2) (55°C for 2 hours) was the fore-runner for both the cultivars. Furthermore, 100 % elimination of Allexivirus, Onion Yellow Dwarf Virus, and Leek Yellow Stripe Virus was observed in T2(HA2) (55°C for 2 hours), as confirmed by RT-PCR. T5(HA5) (60°C for 30 minutes) and T6HW1 (55°C for 10 minutes) though showed complete elimination of Allexivirus and Onion Yellow Dwarf Virus, but failed to eliminate Leek Yellow Stripe Virus. Consequently, treating seed stocks with hot air at 55°C for two hours after every three-four years and practicing planting of large and medium-sized cloves will be instrumental in maintaining the supply and demand of garlic by producing better yields of virus-free and high-quality bulbs.ThesisItem Restricted Management of phosphorus fertilization for tuber yield and quality of potato (Solanum tuberosum L.) genotypes(Punjab Agricultural University, 2023) Vipanpreet Kaur; Navjot SinghThe present study "Management of phosphorus fertilization for tuber yield and quality of potato (Solanum tuberosum L.) genotypes" was conducted at Vegetable Research Farm, Punjab Agricultural University, Ludhiana, and Vegetable Research Farm, Khanaura (Hoshiarpur), during 2021-22. The experiment was conducted in a split-split plot design consisting of two genotypes i.e., Punjab Potato-102 and Kufri Pukhraj in main plots, and two sources of fertilizer i.e., Diammonium phosphate (DAP) and Single Superphosphate (SSP) in sub-plots, and four level of Phosphorus fertilizers i.e., 75, 100, 125 and 150 per cent of recommended dose of Phosphorus fertilizer in sub-sub plot, with three replications. Phosphorus fertilization with DAP @ 93.75 kg/ha (150% of the recommended dose) resulted in the highest emergence percentage, number of shoots, foliage weight, number of tubers per plant, and average tuber weight per plant. Highest tuber yield per hectare was recorded in case of both varieties after 70, 90, and 110 days of harvesting at both locations, with a higher proportion of B-grade tubers followed by C, A, and D grade during all the intervals of harvesting. The tuber quality traits, i.e., dry matter, reducing sugar, total phenols, free amino acids, and total glycoalkaloids improved with the application of a higher dose (150% of phosphorus) of DAP fertilizer. The concentration of nutrient (N, P, and K) uptake was observed to be highest in the root, shoot, and tuber with the application of a higher dose of DAP after 70, 90, and 110 days of harvesting at both locations and varieties. The concentration of nutrient uptake was highest at Khanaura (Hoshiarpur) as compared to Ludhiana. The study on effect of Nano-P fertilizer on tuber yield and quality was conducted at Vegetable Research Farm, Khanaura (Hoshiarpur). The experiment was conducted in a split plot design with two genotypes in main plots, and four levels of Phosphorus fertilizers i.e., 75 per cent, 75 per cent + foliar application of nano-P, 100 per cent, and 100 per cent + foliar application of nano-P in sub plot with three replications. Phosphorus fertilization with @ 62.5 kg/ha (100% of the recommended dose) + foliar application of nano-P resulted in the highest emergence percentage, number of shoots, foliage weight, number of tubers per plant, and average tuber weight per plant to achieve the highest tuber yield per hectare in the case of both varieties after 70, 90, and 110 days of harvesting. The proportion of B-grade tubers was highest followed by C, A, and D grade during all the intervals of harvesting. The tuber quality traits improved with the application of a higher dose (100% of phosphorus + nano-P) of P fertilizer. The concentration of nutrient (N, P, and K) uptake was also highest in the shoot and root with the application of a 100 per cent P + foliar application of nano-P after 70, 90, and 110 days of harvesting in both varieties. The outcome of the present study elucidates the importance of taking an integrated approach to improve the phosphorous utilization by potato crop. This in turn will be useful for management programmes aimed at improving phosphorus use efficiency by adjusting phosphorus fertilization according to location, method of application, cultivar, and source of fertilizers.ThesisItem Embargo Production and Characterization of Haploids in Brinjal (Solanum melongena L.)(Punjab Agricultural University, 2022) Jasmeen Kaur; Sidhu, Mohinder KaurA study was conducted for production and characterization of haploids in brinjal. Induction of haploids in brinjal was attempted through two methods androgenesis and interspecific hybridization. For microspore androgenesis among donor hybrids, PSB-7-2 X P-219 had the highest (14.6 in 63 days) number of calli per ml of microspore suspension, followed by SR-5 x SR-9322 (4.4in 69 days) and SC-15-2 x MR-319 (2.1 in 73 days). Overall, NLN medium supplemented with 2 mg/l 2, 4-D + 20g/l Sucrose induced the maximum number of calli per ml of suspension (13.3 calli in 62 days). In this study, four days temperature pretreatment (35̊ C) in distilled water induced the uppermost number of calli per ml of suspension (11.2 calli in 66 days). The embryogenesis (%) of microspore derived calli was the most effective in MS + 4 mg/l Zeatin + 0.2 mg/l IAA+ 20 g/l of sucrose + 8g/l agar (R1) as compared to MS + 2 mg/l Zeatin + 0.2 mg/l IAA+ 20 g/l of sucrose + 8g/l agar (R2) for all the donor hybrids. In total, 260 plants were rooted on half-strength MS medium and only 65 (25.0 %) could be established as complete plants after hardening. In the confirmation, the molecular analysis with three SSR markers established complete homozygosity (microspore origin) in all the plants developed from two donors. Chromosome counting (n=12) at diakinensis or metaphase stage of meiosis in pollen mother cells categorized 65 regenerants into 5 haploids, 30 doubled-haploids, 29 mixoploids, and one triploid. Many chromosomal abnormalities in PMCs also affcetd the reproductive features of heteroploids. In anter androgenesis of donor hybrids, SC-15-2 x MR-319 and SR-5 x SR-9322 were at par with each other for percentage of calli formation from expanded anthers with values 73.4 % and 71.3 % respectively. Donor hybrid SR-5 x SR-9322 also showed direct embryo regeneration from expanded anthers with value 2.8 %. Among different media combinations, the maximum calli formation from expanded anthers with in minimum days was observed in MS + 2 mg/l IAA + 0.5 mg/l Kinetin (65.8 % and 28.5 days). Direct embryo regeneration from expanded anthers was observed only in media composition MS + 2 mg/l IAA + 0.5 mg/l Kinetin (1.6 %). However, the highest calli regeneration was observed in donor hybrid SR-5 x SR-9322 (15.5%) with in minimum days (21.7 days). Among media combinations, MS + 0.2 mg/l IAA + 4 mg/l Zeatin (19.5 %) had the maximum calli regeneration in the minimum number of days (21.4 days). Plants obtained through anther androgenesis via direct embryos regeneration from expanded anthers in SR-5 x SR-9322 at media combination MS + 2 mg/l IAA + 0.5 mg/l Kinetin also confirmed their genetic origin. Morphological observations and molecular marker EEMS 37 confirmed the possibility of the haploid nature of these plants. In wide hybridization, no chromosome elimination was observed in plants. In conclusion, microspore culture of F1 donor was the best approach to develop homozygous lines of genetic origin and considerable morphological and genetic variability.
