ThesisItem Open AccessIdentification for novel alleles for acidity tolerance in rice(College of Post Graduate Studies in Agricultural Sciences, CAU-Imphal, Umiam, 2015-10) Sunderi, Julia Yumnam; Tyagi, WrichaProblem soils comprise a considerable proportion of rice production of rice production areas, which are invariably associated with poverty due to low and unstable productivity. Majority of these soils are concentrated in North-Eastern region of India, with nearly 65 % of its area being under extreme forms of soil acidity (pH below 5.5). Under acidic soil conditions phosphorus (P) becomes unavailable to the crop plants coupled with aluminium and iron toxicity. Screening of Indian germplasm has been initiated for P deficiency tolerance but performance of these genotypes in acidic soils is not properly documented. Therefore, it necessitates the use of different genotypes present in this region along with reported tolerant / susceptible checks for further molecular breeding programmes. Hydroponics based screening was done to identify low P deficiency and aluminium and iron toxicity tolerant genotypes. Rice seedlings screened under low P condition for 15 and 25 days revealed LR 23 and LR 26 as tolerant genotypes. Genotypes like UR 5 and UR 29 were identified as tolerant when screened under 0.54 mM of Al3+for 5 days based on basis of percentage increase in root and shoot biomass. Pot experiment conducted for 25 days for aluminium toxicity tolerance (0.54 mM of Al3+) led to identification of LR 39, LR 56 and LR 13 as tolerant genotypes. Field screening was done for P deficiency and iron toxicity tolerance. LR11, LR 15, LR 18 and LR 26 were tolerant to iron toxicity both under hydr oponics and field conditions. Rice genotypes LR 23 and UR 95 showed high P uptake and PUE (phosphorus use efficiency but P efficiency was highest in LR 56. Lowland field screening revealed that total dry weight was positively correlated with iron content i n the shoots. PUE was also positively correlated with phosphorus uptake and plant dry weight. Different mechanism dealing with excess iron was found in rice genotypes evaluated in the present study. Allele mining across OsPSTOL 1 and Dirigent-like gene (Os Pup K 20-2) was done for P deficiency tolerance. Nucleotide substitutions found in LR 39 and UR 29 for OsPSTOL 1 was similar with O. glaberrima. LR 39 and UR 29 showed mixed haplotypes different from those reported previously for tolerant genotypes like Kasalath and O. glaberrima. Our results indicate that new markers for molecular breeding will have to be designed for this novel haplotype. One novel allele was also found in the 3 ′ UTR of OsPSTOL 1 for LR 39 and UR 29. Allele mining across Dirigent- like gene revealed Kasalath specific 3 bp deletion at position 665 bp in LR 11, LR 39 and LR 55. In addition to that, four novel alleles were also found. Sequencing across LOC_Os03g28250 (novel gene for P tolerance) revealed change in the protein sequence due to 8 bp deletion. A total of 15 novel alleles were found in the coding region of LOC_Os03g28250. A 45 bp deletion was observed at 572 bp of the genomic region. The insilco analyses of novel low P induced transcripts suggest that out of nine transcripts, four were associated with other abiotic stress responses. The upstream region of all the novel transcripts carries a common motif EBOXBNNAPA (CCGAC). LTRE motif was present in LOC_Os 04g26330, LOC_Os02g03640, LOC_Os03g28250, LOC_Os01g35830 and LOC_Os03g32330. Coordinated expression revealed that the transcripts LOC_Os01g37260, LOC_Os01g35830 and LOC_Os04g10214 were over expressed in roots of arsenic sensitive variety Azucena. Allele mining of LOC_ Os02g03900 identified one putative polymorphism in aluminium sensitive genotype, LR 55. In presence of aluminium, the expression of LOC_Os07g34520 was upregulated in LR 55. On the contrary, expression of LOC_Os02g03900 was upregulated in aluminium treated roots in UR 29 and downregulated in UR 5. An on- synonymous substitution was found in at position 2241 for OsFRLD 1 gene which led to the change of valine (GTA) to leucine (CTA). Four biparental crosses derived from diverse parents (with respect to the allelic status of OsPSTOL 1, Dirigent-like gene, OsFRLD 1 and iron uptake) were evaluated in the lowland acidic field condition. The cross between LR 15 and LR 18 performed better than the other three crosses based on the yield parameter. The cross between LR 11 and LR 26 can be used for mapping under low P while for iron toxicity cross between LR 13 and LR 26 can be used. The cross LR 11 and LR 26 was selected for mapping in low P condition. Total dry weight was used as a trait for bulk segregant analysis which revealed three informative markers. Iron toxicity screening for F3 population (LR 26 X LR 13) revealed that number of crown root was negatively correlated with root length. This data along with screening data suggests that the number of crown roots, root length and leaf bronzing can be taken as traits for consideration for identifying iron toxicity tolerance rice genotypes.