QTL Mapping And Genetic Dissection Of Strong Seed Dormancy In N22 (Oryza Sativa L.)

Grain dormancy is an important trait for breeders in many cereals because of its association with preharvest sprouting, which can damage end-use quality such as seed quality and yield in rice. In conventional rice breeding, high-yield varieties are liable to initiate germination before harvest given suitable environmental conditions around the time of crop maturity in recent years, strongly influence quality of rice. Strong levels of seed dormancy are correlated with a low probability of PHS and vice versa. Excessive dormancy of course can also be problematical, because it leads to uneven seedling establishment. Therefore, breeding for an intermediate level of dormancy in rice is highly desirable. The indica cultivar N22 has very strong level of dormancy. Here, two different strategies were used to genetic analysis of N22. The first, the advanced backcross (AB) populations and near-isogenic lines (NILs) contained qSdn-1 or qSdn-5 that were two major effect dormancy QTL in N22 was developed respectivily. qSdn-1 and qSdn-5 was fine mapping in a narrow region using AB-population and the effect of these two locus were also verfied using NILs. The second, the seeds of N22 were treated with 400Gy 60Co gamma-radiation, the mutants associated with seed dormancy were screened, and the simple genetic analyses were done for these mutants. These two strategies will provide some useful information to reveal the genetic mechanism for seed dormancy of N22.1. The intrachromosomal positions of the two grain dormancy quantitative trait locus (QTL) qSdn-1 (chromosome 1) and qSdn-5 (chromosome 5) were obtained from the segregation analysis of the advanced backcross populations derived from the cross between rice cultivars N22 and Nanjing35. Marker-assisted selection (MAS) was applied to select derivatives carrying one or both of qSdn-1 and qSdn-5 in a genetic background which was nearly isogenic to Nanjing35.In 2008, an analysis of dormancy in the BC4F2 population allowed qSdn-1 to be located between the simple sequence repeat (SSR) markers RM11669 and RM1216; the QTL explained 24.58% of the overall phenotypic variation and the most closely linked marker was RM11694. qSdn-5 was mapped between RM480 and RM3664, and explained 17.58% of the overall phenotypic variation. The SSR locus RM19080 mapped within 0.4 cM of qSdn-5. No epistasis was observed between qSdn-1 and qSdn-5. The mean germination rates of lines containing qSdn-1, qSdn-5 and both qSdn-1 and qSdn-5 was 7.9, 11.1 and 6.1%, respectively, whereas that of the check line lacking both QTL was 86.3%. The dormancy of both qSdn-1 and qSdn-5 could be readily broken by a 7-day post-harvest treatment at 50℃.Later, qSdn-1 and qSdn-5 were fine mapped using advanced backcross populations BC5F2 and BC5F3 in 2009 and 2010. In 2009,95 extreme recombinant plants were identfied using the SSR markers RM128 and RM11781 from 7300 BC5F2 (qSdn-1), the phenotypes of these recombinant plants were verified by the progenies (BC5F3) in 2010, qSdn-1 was mapped between SSR marker L24 and L34 with about 655kb, co-segregating with L27; using the same method,111 extreme recombinant plants were identfied using RM7452 and RM3664 from 5888 BC5F2 (qSdn-5), finally, qSdn-5 was mapped between Indel marker 15-2 and SSR marker RM19080 with 122kb, co-segregating with Indel marker 15-6. Fine mapping of qSdn-1 and qSdn-5 established good base for map-based cloning of these two QTL. The SSR loci linked most tightly to qSdn-1 and qSdn-5 are suitable for MAS for reduced pre-harvest sprouting in rice.When we fine mapped qSdn-1 and qSdn-5, three NILs, NIL (qSdn-1), NIL (qSdn-5) and NIL (CK) were developed using phenotype and marker-assisted selection. The germination rates of NIL (qSdn-1), NIL (qSdn-5) and NIL (CK) were 23%,35% and 98%, respectively in 2010, while the major agronomic traits of the NILs were same with Nanjing35. This also veried qSdn-1 and qSdn-5 act very important effect for seed dormancy of N22.Seed dormancy have a tight connection with plant hormones, the seed germinations of NIL (qSdn-1) and NIL (qSdn-5) were treated with hormone and adversity stress, the results showed different sensitivity to ABA, GA and NaCl between NIL (qSdn-1) and NIL (qSdn-5), NIL (qSdn-5) displayed more sensitivity than NIL (qSdn-1), there were no sensibly different between NIL(qSdn-1) and NIL (qSdn-5) when treated with IAA, the different sensitivity indicated the genetic mechanism of the gene underlying qSdn-1 and qSdn-5 is different.2. Two weak dormancy mutants, designated Q4359 and Q4646, were obtained from the rice cultivar N22 after treatment with 400Gy 60C0 gamma-radiation. The germination rates of Q4359 and Q4646 were 43% and 45%, respectively after 35d heading, higher than <2% of wide-type N22. The dormancy of the mutant seeds was more readily broken when exposed to period of room temperature storage. The mutants also showed a reduced level of sensitivity to ABA and NaCl compared to the N22 cultivar, although Q4359 was more insensitive than Q4646. The germination rate of N22 increased with accruement of GA concentration, while Q4359 and Q4646 displayed insensitive to GA. The germination rates of N22, Q4359 and Q4646 all increased with accruement of IAA concentration, no obvious different.A genetic analysis indicated that in both mutants, the reduced dormancy trait was caused by a single recessive allele of a nuclear gene, but that the mutated locus was different in each case. There is one or two major gene(s) in N22, in previous study, qSdn-1 and qSdn-5 were detected as major effect QTL. Whether or not the reduced dormancy accociated with these two QTL? So we detected the dormancy QTL of Q4359 and Q4646 using two segregation populations, Q4359/Nanjing35 F2 and Q4646/Nanjing35 F2. In Q4359/Nanjing35 F2,3 QTL, qSdNj-3, qSdn-5, and qSdn-9 were detected on chromosome 3,5 and 9, respectively. The QTL qSdn-9 was determined to be a novel dormancy locus, and it was mapped between SSR markers RM7038 and RM105 with a LOD score of 5.5, explaining 11.5% of the overall trait variation. The major dormancy QTL qSdn-1 was not detected in Q4359/Nanjing35 F2. Two QTL, qSdn-1 and qSdNj-3, were detected in Q4646/Nanjing35 F2, the position of qSdNj-3 was accorded with the QTL in Q4359/Nanjing35 F2, and qSdn-5 was not detected in Q4646/Nanjing35 F2. The following QTL:qSdn-2, qSdn-7 and qSdn-11 were not detected in the two populations. Therefore, qSdn-1 and qSdn-5 could be inherited as the major dormancy locus, but qSdn-5 in Q4359 and qSdn-1 in Q4646 were not detected. Whether the reduced dormancy phenotype caused by qSdn-1 and qSdn-5 mutated or not will require further investigation.