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2022 - 20243
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Subject: Vegetable Science × Start date: 2022 × Thesis Type: Ph.D ×
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    Development of monoecious lines in muskmelon (Cucumis melo L.) through marker assisted selection
    (Punjab Agricultural University, 2024) Nidhi; Sharma, Sat Pal
    In 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.
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    Inheritance and molecular mapping of gene(s) associated with yellow vein mosaic virus disease (YVMVD) resistance in pumpkin (Cucurbita moschata Duch. ex Poir)
    (Punjab Agricultural University, Ludhiana, 2022) Verma, Neha; Dhatt, Ajmer Singh
    Worldwide begomoviruses causative diseases are major reason of quantitative and qualitative losses in pumpkin. The three types of viral symptoms were molecularly characterized as SqLCCV, ToLCNDV and their mixed infection (MI-Sq/To). Artificial screening, histopathological study and PCR confirmed PVR-1343 as a novel resistant source against both begomoviruses. To understand the inheritance and mapping of begomovirus resistant gene(s), segregating populations between PVR-1343 and five susceptible lines (P-135, P-6711, Punjab Samrat, MBN-6242 and P-278) were developed and artificially inoculated with Bemisia tabaci Asia II strain against both viruses and their mixed infection in enclosed structures for three consecutive years (2018-2020). Disease scoring at 75DAS rendered all F1 progenies as susceptible indicating recessive nature of resistance with no maternal or cytoplasmic effects. Chi-square analysis showed best fit of 9(S):3(MS):3(MR):1(R) and 1(S):1(MS):1(MR):1(R) ratio in F2 and backcross with resistant parent respectively. Phenotypic segregation ratio of F2:3 confirmed the F2 genotypic ratio of 1:2:2:4:1:2:1:2:1. This substantiated that multiple begomovirus resistance in PVR-1343 is controlled by a digenic recessive gene. Moreover, F2:3 progenies resistant to SqLCCV were also resistant to ToLCNDV and MI-Sq/To and vice versa, suggesting that common or tightly linked gene(s) are involved in resistance to both viruses. To map these genes, WGRS coupled with QTLseq of two extreme bulks (resistant and susceptible) of cross PVR-1343×P-135 along with parents were employed and two genomic regions viz., qMI-Sq/To7.1 on chromosome 7 (3.18 to 4.70Mb) and qMI-Sq/To17.1 on chromosome 17 (7.30 to 8.17Mb) were identified. Nine polymorphic SNPs identified within the highly significant qMI-Sq/To7.1 region were converted into KASP marker. The KASP genotyping of 177 F2 individuals narrowed down the qMI-Sq/To7.1 interval to a 103Kb region flanked by Cmo3914729 and Cmo4018182 KASP markers. One of the flanking SNP (Cmo4018182 KASP marker) imparting resistance against MI-Sq/To, accurately predicted the disease reaction of 91 per cent of diverse Cucurbita genotypes. This SNP showed nonsynonym substitution in the coding region of putative candidate syntaxin-121 protein and might be responsible for diminishing or abolishing the viral infections. This study has thus, seeded the path for marker assisted selection for efficient introgression of resistance into elite C. moschata lines and providing a viewpoint for gene cloning.
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    Generation of frost tolerant potato (Solanum tuberosum L.) clones through intervarietal hybridization and expression analysis of putative frost tolerance genes
    (Punjab Agricultural University, Ludhiana, 2022) Angmo, Dechen; Sharma, Sat Pal
    The present study was carried out with an overall aim to generate frost tolerant potato clones and optimization of phenotyping techniques in order to complement the natural field selection criteria. The physiological responses to low temperature stress were explored using four potato genotypes viz. Kufri Anand, J-2/19, MS/7-645 and Kufri Pukhraj under in vitro controlled conditions. Significant variation was observed amongst genotypes for parameters associated with frost tolerance in potato such as cell membrane stability, activity of PSII, photosynthetic efficiency and leaf morpho-anatomical traits. The genotypes J/2-19 and Kufri Anand exhibited higher PN, gs and ΔF/Fm’values and lower electrolyte leakage as compared to the sensitive genotype Kufri Pukhraj upon cold acclimation and during recovery days after freezing exposure. Similarly, J-2/19 genotype also recorded higher adaxial trichome density than the sensitive genotype MS/7-645. In order to identify the fold changes in the expression of three candidate genes involved in cold stress responses in potato viz. dehydrin, hsp70 and SAD a qRT-PCR study was also performed. Overall an increase in gene expression was observed in cold acclimated plants of all the genotypes compared to non-acclimated plants. However, the increase was more pronounced in tolerant genotype (J-2/19) as compared with susceptible genotypes (MS/7-645 and Kufri Pukhraj). In the summer 2018, the segregating progenies were developed through hybridization among contrasting parents viz. J-2/19  MS/7-645 (PAU3), MS/7-645  J-2/19(PAU7) and MS/7-645  CP-3765 (PAU9) at Keylong, HP. In F1C1 and F1C2 generations, the characterization of the generated potato clones was accomplished on the basis of leaf morpho-anatomical traits and electrolyte leakage assay. To determine the association between the key traits and potential frost tolerant potato clones, various statistical approaches, such as cluster analysis, principal component analysis and genotype by trait biplot analysis were performed. Total 11, 10 and 8 clones from PAU3, PAU7 and PAU9 crosses, respectively were identified exhibiting potential frost tolerant characteristics with desirable horticultural traits for their further evaluation. Thus, the findings of this study concluded that in addition to electrolyte leakage the chlorophyll fluorescence and leaf gas exchange parameters can be effectively utilized for screening frost tolerant potato genotypes, but with certain limitations. Furthermore, the leaf morpho-anatomical traits have been confirmed to be associated with frost tolerance through decrease in foliage damage and electrolyte leakage. Therefore, these traits can potentially be used for screening of large segregating populations for freezing tolerance in the early clonal generations in potato.