Mapping Of A Novel Dwarfing Rice And Expression Analysis Of Gibberellin Key Metabolic Enzyme Genes

Breakthrough of crop breeding depends on the discovery and use of key gene resources. Identification and exploration of new dwarfing gene is significant to rice breeding and relative research on dwarfing mechanism. The plant material was an especially dwarfing rice derived from spontaneously mutation (named'Teaidao-2'). We characterized sd-1 locus of'Teaidao-2'by PCR amplification with the primers which were designed by ourselves, the result demonstrated that'Teaidao-2'might mutated from parents with sd-1 locus. Physiological analysis revealed that the GA signaling pathway of'Teaidao-2'was normal and exogenous GA3 could not rescue the dwarfing phenotype of'Teaidao-2', suggested that gibberellin was not involved in the dwarfing phenotype. Genetic analysis carried out on F1 and F2 population from the cross between'Teaidao-2'and'Nipponbare'showed that the height of F1 population was normal, the ratio between dwarfing plants and normal plants was 1:3 in F2 population, so the dwarfing phenotype of'Teaidao-2'was regulated by a single recessive gene. The dwarfing gene was referred to as ED, a F2 population derived from cross between'Teaidao-2'and'Nipponbare'was constructed and 243 pairs of simple sequence repeat (SSR) markers were used to map the ED locus. With the aid of Mapmaker/EXP Version 3.0 and SSR marker analyses, the ED gene was located on chromosome 12, flanked by SSR markers RM519 and RM235 with the genetic distances of 15.9 and 22.0 cM, respectively. The other linked markers included RM101 and RM17 with the genetic distances of 29.0 and 27.3 cM. These linked markers were in the order of RM101-RM519-ED-RM235-RM17.The GAs form a large family of tetracyclic diterpenoid phytohormones that are involved in the regulation of various growth and developmental processes in higher plants, including seed germination, stem elongation, flowering and fruit development. Particularly, the role of GA in plant growth is represented by the promotion of stem elongation since almost all of the GA-deficient mutants exhibit dwarfism. In order to find out the mutational mechanism of'Teaidao-2', we measured the expression of gibberellin key metabolic enzyme genes. According to the cDNA sequences of rice gibberellin key metabolic enzyme genes from GenBank, primers were designed in conserved regions.Firstly, we adopted traditional RT-PCR to detect the transcription level of gibberellin key metabolic enzyme genes of'Teqing'and'Teaidao-2'. Meanwhile, with optimizing of reaction condition, we established a real-time quantitative PCR detection method by standard curve analysis. The assays demonstrated that the specification of primers was strong and the correlatability of standard curves was fine, the experiment had a high credibility, and could be promoted to the research of rice GA relative mutants.Detection was carried out on'Teqing'and'Teaidao-2'for a more exact expression level. The results demonstrated that the gibberellin 20-oxidase genes (GA20ox2) in both'Teqing'and'Teaidao-2'were transcripted at low level, which was in accordance with the conclusion of sd-1 locus characterization, and the rest of gibberellin key metabolic enzyme genes of'Teqing'and'Teaidao-2'were expressed at normal level, furtherly clarified gibberellin would have no relation with the especial dwarfism of'Teaidao-2'. The same detection was also be taken on'Zhong 9B'and'SV 9B', which was the dwarf mutant of'Zhong 9B'from somatic mutagenesis. The results indicated that the ent-kaurene oxidase gene 2(KO2) and gibberellin 3-oxidase gene 2(GA3ox2) were down-regulated apparently in'SV 9B', which might be relevant to the mutation of dwarfism.