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ThesisItem Open Access Genetics of shattering resistance in rice (Oryza sativa L.)(Department of Plant Breeding and Genetics, College of Horticulture, Vellanikkara, 2020) Akhil, K P; KAU; Biju, SRice is the staple food crop of Kerala. Among the plethora of rice varieties available Jyothi is by far the most popular in the state owing to its consumer preference. Easy seed shattering leads to significant loss in Jyothi which can be overcome by transferring shattering resistance to it. Screening of genotypes for shattering resistance identified donors like Athira and Triveni with good combining ability. The present experiment was thus envisaged to transfer shattering resistance to Jyothi from the donors and to study the nature of inheritance of seed shattering trait using six parameter model of generation mean analysis. Observations were recorded in the two crosses viz., Jyothi x Aathira and Jyothi x Triveni for eleven quantitative characters viz. days to 50 per cent flowering, plant height, tillers per plant, panicles per plant, panicle length, spikelets per panicle, seeds per panicle, days to maturity, test weight, grain yield per plant and shattering per cent in six generation (P1, P2, F1, F2, B1 and B2) obtained from the two crosses. Panicle per plant, test weight, grain yield per plant and shattering per cent recorded significant heterosis in the desirable direction in both the crosses. Mean effect (m) was significant and positive for all the characters studied in both the crosses. The additive component (d) was significant and positive for days to 50 per cent flowering, tillers per plant, panicles per plant, days to maturity, grain yield per plant and shattering per cent in the cross Jyothi x Triveni while the other characters recorded significant and negative additive gene effect. The characters tillers per plant, test weight and shattering per cent showed significant positive additive gene effect in the cross Jyothi x Aathira and rest of the characters showed significant and negative additive component. Dominance gene effect was observed to be significant and negative for most of the characters studied in both the crosses. Non-allelic interactions were observed to be significant in most of the characters except days to 50 per cent flowering, tillers per plant, days to maturity and shattering per cent in the cross Jyothi x Aathira. The epistatic interaction model of generation mean analysis was found adequate for obtaining gene actions for all the characters. Among the three type of interactions dominance x dominance interaction was found to be more important for all the characters. Additive x additive and additive x dominance gene interactions were found to be equally important for most of the characters studied in both the crosses. Dominance gene effect was observed controlling the genetic variance in most of the traits studied. Presence of additive and non-additive along with epistatic interaction revealed the complex nature of inheritance of the characters. Predominant dominant gene effect along with duplicate epistasis limits the scope of direct selection. In the F2 populations characters showed high heritability in both the populations studied. Plant height, panicle length and days to maturity were found to have low genetic advance. Plant height recorded moderate genetic advance, whereas, other characters exhibited high genetic advance in F2 population derived from the cross Jyothi x Triveni. Days to maturity showed low genetic advance, days to 50 per cent flowering and panicle length recorded moderate genetic advance and other characters recorded high genetic advance in F2 population derived from the cross Jyothi x Aathira. As shattering resistance is governed predominantly by additive gene effects selection in the early segregating generations will be highly rewarding. The selected lines may be advanced to further generations to identify lines with high yield coupled with shattering resistance and can be forwarded to develop high yielding varieties with low shattering or bi-parental mating among the selected lines may be undertaken to recover superior recombinants with high yield and shattering resistance.ThesisItem Open Access Development of doubled haploids for iron toxicity tolerance in rice (Oryza sativa L.)(Department of Plant Breeding and Genetics, College of Horticulture, Vellanikkara, 2020) Chakravarthi, Marri; KAU; Dijee, BastianRice is the staple food crop of the people of Kerala. The production of rice in Kerala is impeded by many biotic and abiotic stresses. Iron toxicity is one of the major abiotic stresses of acidic soils in Kerala. The present study “Development of doubled haploids for iron toxicity tolerance in rice (Oryza sativa L.)” was taken up with the objective of developing doubled haploids with tolerance to iron toxicity through in vitro anther culture. Doubled haploids being homozygous stable lines, can be used both as immortal populations for QTL mapping as well as varieties, if suitable agronomic traits are present in them. Therefore, this study was taken up which comprised of three experiments. In experiment I, two tolerant genotypes (Swetha and Mangala Mahsuri) and two susceptible genotypes (Cul-90-03 and Cul 8709) were crossed with Tulasi. In the second experiment the parents along with their hybrids were evaluated for biometric characters and yield. The hybrids obtained from the crosses were significantly different for all the characters observed. Panicles per plant and grain yield per plant recorded highly significant and positive relative heterosis and heterobeltiosis. The anthers from the F1s produced in the previous experiment were used for anther culture studies in the third experiment. Sterilisation with various sterilants like 70 % ethanol, 0.1 % mercuric chloride, 5.25% sodium hypochlorite and their combinations were done. Explant sterilisation with 5.25% sodium hypochlorite for 5 to 20 minutes was effective in controlling contamination in the in vitro cultures. Study to know the best number of days of cold pre-treatment at 10oC was done (0, 3, 6, 9, 12, 15 days). Cold pre-treatment of anthers at 10oC for 9 days was found to be optimum for most of the genotypes studied. Two different media viz., N6 and B5 were tried with different combinations of 2,4 - D and Kn. N6 media responded better than B5 media for callus induction in all the growth hormone combinations. The effect of carbon source on callusing was studied using maltose and sucrose at various levels (30,40,50 mg/L) and it was found that maltose at 30g/L gave the best callus induction (7.95%). Among the auxins used in the present study,2,4-D was found to be better than NAA for callus induction while among cytokinins, Kinetin responded better than BAP for callus induction. The growth hormone combination 2,4- D (2mg/L) + Kn (0.5mg/L) was adjudged the best for callus induction. Additives like silver nitrate (AgNO3), casein hydrolysate (CH), yeast extract (YE), proline and activated charcoal were added to the basal media to improve callusing. when AgNO3 is applied from 0 to 1ml with 0.1 ml gradation, 0.5-0.6ml of 0.1N AgNO3 was found to be better in callus induction as well as the days to callus induction was reduced at these concentrations. 250 – 500 mg/L of CH and 250mg/L proline were found to induce significant response for callusing while activated charcoal and yeast extract did not have any considerable effect on callus induction. Hybrid H1 recorded good callus response when maltose at three levels was tried in the media (30, 40, 50g/L) whereas hybrid H2 gave uniform response at all the levels of maltose. There was significant variation among the genotypes in their response to all the factors studied. The best responses were as follows: - H1(Swetha x Tulasi) - N6+2,4-D (2mg/L) + Kn (0.5mg/L) + 30g/L maltose +0.5ml (0.1 N AgNO3) + 250mg/L proline + 250mg/L CH +2.5g/L gelrite gellan gum. H2(Mangala Mahsuri x Tulasi) - N6+2,4-D (2mg/L) +Kn (0.5mg/L) + 30g/L maltose +0.5ml (0.1N AgNO3) +250mg/L proline+250mg/L CH +2.5g/L gelrite gellan gum. H3(Cul-90-03 x Tulasi) - N6+2,4-D (2mg/L) +Kn (0.5mg/L) + 30g/L maltose + 0.5ml (AgNO3) + 250mg/L proline + 250mg/L CH + 2.5g/L gelrite gellan gum. H4(Cul 8709 x Tulasi)- B5 + 2,4-D (2mg/L) + Kn (0.5mg/L) + 30g/L maltose + 0.5ml (0.1N AgNO3) + 250mg/L proline + 250mg/L CH + 2.5g/L gelrite gellan gum. The calli obtained from the different genotypes were plated on two different callus regeneration media R1 (MS+ NAA(1mg/L) + Kn (2mg/L) + IAA (0.5mg/L) + CW (5%)) and R2 (MS+NAA (0.25mg/L) + BAP (0.75mg/L) + Kn(0.25mg/L)). There was no response in R1 media. Callus regenerated into plantlets in R2 but all the plantlets obtained were albinos leading to mortality and hence plantlet hardening and field planting could not be undertaken.ThesisItem Open Access Marker assisted backcross breeding for pyramiding genes conferring resistance to bacterial blight in rice variety uma(Department of Plant Breeding and Genetics, College of Horticulture, Vellanikkara, 2018) Megha, L M; KAU; Rose Mary FranciesAs in other rice growing locales around the world, bacterial blight (BB) caused by Xanthomonas oryzae pv. oryzae (Xoo) assumes a huge role in deciding rice profitability in Kerala. The elite rice varieties of Kerala, both PTB 39 (Jyothi) and Mo 16 (Uma), are found to be extremely susceptible to bacterial blight. Since both the major rice cropping seasons (virippu and mundakan) in the state coincide with monsoons, the control of the disease through chemicals or biological agents proves inadequate owing to the washing-off of the applied materials. Host-plant resistance is advocated as the most effective breeding strategy to combat the bacterial blight (BB) pathogen. Considering the impact of the disease on food security and sustainability, efforts were taken to introgress three R-genes (xa5, xa13 and Xa21) into the variety Uma from donor parent Improved Samba Mahsuri (ISM) through Marker Assisted Selection (MAS). Further, backcrossing to Uma (recurrent parent) and advancing the resultant BC1F1s have resulted in production of BC2F1 generation (21 Nos.). The present study aimed to identify BC2F1 plants pyramided with genes (xa5, xa13 and Xa21) conferring resistance to bacterial blight using functional markers. In addition, advancing the R-gene introgressed BC2F1s to BC3F1 and BC2F2 generation was envisaged. Pathotyping of BC1F2s (850 Nos.) and generating BC1F3s from the plants exhibiting resistance to BB pathogen was also aimed at. Foreground selection of the BC2F1 individuals using the xa5 gene linked STS marker RG 556 and functional marker xa5 SR confirmed the presence of the R-gene in the parents as well as the 21 BC2F1 individuals. Screening of BC2F1 individuals with STS marker RG 136 linked to R-gene xa13 and functional marker xa13 promoter revealed that the BC2F1 Plant No. 8.3.9.10 was heterozygous at xa13 locus while, all other BC2F1 individuals possessed alleles similar to that of the recurrent parent (RP) Uma. The result also pointed out that BC2F1 Plant No. 8.3.9.10 was a 2-R-gene pyramid (xa5xa5 + Xa13xa13). Foreground selection with STS marker pTA 248 to detect the presence of Xa21 gene revealed that none of the BC2F1 plants analysed except BC2F1 Plant No. 8.3.9.10, possessed the resistant allele of R-gene Xa21. However, presence of alleles of both the parents in Plant No. 8.3.9.10 indicated that it was heterozygous at Xa21 locus. Results obtained thus revealed that, of the 21 BC2F1s subjected to foreground selection, BC2F1 Plant No. 8.3.9.10 was the only 3-R-gene introgressed pyramid (xa5xa5 + Xa13xa13 + Xa21xa21). In addition, the 3-R-gene introgressed BC2F1 Plant No. 8.3.9.10 was subjected to background screening using 22 rice microsatellite (RM) markers. Background profiling revealed that the banding pattern in Plant No. 8.3.9.10 was similar to recurrent parent in case of thirteen RM markers. The plant was found to be heterozygous at five other marker loci. Considering the segregation of the 22 markers, the magnitude of recovery of recurrent parent genome in 3-R-gene introgressed BC2F1 Plant No. 8.3.9.10 was found to be 81.82 per cent. The dendrogram based on molecular data grouped the individuals into two major clusters. Cluster 1 was monogenic with only the donor parent ISM and cluster 2 comprised of the recurrent parent Uma and Plant No.8.3.9.10, further suggesting that the 3-R-gene introgressed BC2F1 Plant No. 8.3.9.10 was more similar to the recurrent parent Uma. Wide variability was observed among the BC2F1 individuals for various morphological traits. The 3-R-gene introgressed BC2F1 Plant No. 8.3.9.10 was shorter in duration than both the parents and also possessed red kernels similar to the recurrent parent Uma. The dendrogram generated based on the morphological characters also indicated greater similarity between the 3-R-gene pyramid and recurrent parent Uma. The identified 3-R-gene pyramid (BC2F1 Plant No. 8.3.9.10) was backcrossed to both recurrent parent Uma as well as selfed resulting in production of BC3F1s (15 Nos.) and BC2F2s (28 Nos.) respectively. Bioassay of BC1F2 population (106 Nos.) through leaf clipping method of pathotyping suggested by IRRI (1991) revealed that more than half the BC1F2 individuals screened exhibited resistance to BB pathogen. The plants that exhibited moderate reaction to BB infection ranged between 16.87 per cent and 34.78 per cent in progeny of BC1F2 Plant No. 8.3.2 and BC1F2 Plant No. 8.3.9 respectively. Selfing of the BC1F2 individuals exhibiting resistance and moderate resistance to BB pathogen resulted in production of 725 BC1F3 seeds. Modern molecular techniques make it possible to use markers and probes to track the simultaneous introgression of several R-genes into a single cultivar during a crossing programme. Foreground and background profiling of backcross generations can ensure precise identification of R-gene introgressed genotypes that resemble the recurrent parent Uma.