Molecular dissection of connected rice populations revealed important genomic regions for agronomic and biofortification traits.

Breeding staple crops with increased micronutrient concentration is a sustainable approach to address micronutrient malnutrition. We carried out Multi-Cross QTL analysis and Inclusive Composite Interval Mapping for 11 agronomic, yield and biofortification traits using four connected RILs populations of rice. Overall, MC-156 QTLs were detected for agronomic (115) and biofortification (41) traits, which were higher in number but smaller in effects compared to single population analysis. The MC-QTL analysis was able to detect important QTLs viz: qZn5.2, qFe7.1, qGY10.1, qDF7.1, qPH1.1, qNT4.1, qPT4.1, qPL1.2, qTGW5.1, qGL3.1 , and qGW6.1 , which can be used in rice genomics assisted breeding. A major QTL (qZn5.2 ) for grain Zn concentration has been detected on chromosome 5 that accounted for 13% of R2. In all, 26 QTL clusters were identified on different chromosomes. qPH6.1 epistatically interacted with qZn5.1 and qGY6.2 . Most of QTLs were co-located with functionally related candidate genes indicating the accuracy of QTL mapping. The genomic region of qZn5.2 was co-located with putative genes such as OsZIP5, OsZIP9, and LOC_OS05G40490 that are involved in Zn uptake. These genes included polymorphic functional SNPs, and their promoter regions were enriched with cis-regulatory elements involved in plant growth and development, and biotic and abiotic stress tolerance. Major effect QTL identified for biofortification and agronomic traits can be utilized in breeding for Zn biofortified rice varieties.

Meta-QTL s and haplotypes for efficient zinc biofortification of rice.

Biofortification of rice with improved grain zinc (Zn) content is the most sustainable and cost-effective approach to address Zn malnutrition in Asia. Genomics-assisted breeding using precise and consistent Zn quantitative trait loci (QTLs), genes, and haplotypes can fast-track the development of Zn biofortified rice varieties. We conducted the meta-analysis of 155 Zn QTLs reported from 26 different studies. Results revealed 57 meta-QTLs with a significant reduction of 63.2% and 80% in the number and confidence interval of the Zn QTLs, respectively. Meta-quantitative trait loci (MQTLs) regions were found to be enriched with diverse metal homeostasis genes; at least 11 MQTLs were colocated with 20 known major genes involved in the production of root exudates, metal uptake, transport, partitioning, and loading into grains in rice. These genes were differentially expressed in vegetative and reproductive tissues, and a complex web of interactions were observed among them. We identified superior haplotypes and their combinations for nine candidate genes (CGs), and the frequency and allelic effects of superior haplotypes varied in different subgroups. The precise MQTLs with high phenotypic variance, CGs, and superior haplotypes identified in our study are useful for an efficient Zn biofortification of rice and to ensure Zn as an essential component of all the future rice varieties through mainstreaming of Zn breeding.

Introgression of tsv1 improves tungro disease resistance of a rice variety BRRI dhan71.

Rice Tungro disease poses a threat to rice production in Asia. Marker assisted backcross breeding is the most feasible approach to address the tungro disease. We targeted to introgress tungro resistance locus tsv1 from Matatag 1 into a popular but tungro susceptible rice variety of Bangladesh, BRRI dhan71. The tsv1 locus was traced using two tightly linked markers RM336 and RM21801, and background genotyping was carried out using 7 K SNPs. A series of three back crosses followed by selfing resulted in identification of plants similar to BRRI dhan71. The background recovery varied at 91-95% with most of the lines having 95%. The disease screening of the lines showed moderate to high level of tungro resistance with a disease index score of ≤ 5. Introgression Lines (ILs) had medium slender grain type, and head rice recovery (59.2%), amylose content (20.1%), gel consistency (40.1 mm) and gelatinization temperature were within the acceptable range. AMMI and Kang's stability analysis based on multi-location data revealed that multiple selected ILs outperformed BRRI dhan71 across the locations. IR144480-2-2-5, IR144483-1-2-4, IR144484-1-2-2 and IR144484-1-2-5 are the most promising lines. These lines will be further evaluated and nominated for varietal testing in Bangladesh.

Genetic Analysis of Agronomic Traits and Grain Iron and Zinc Concentrations in a Doubled Haploid Population of Rice (Oryza sativa L.).

The development of micronutrient dense rice varieties with good agronomic traits is one of the sustainable and cost-effective approaches for reducing malnutrition. Identification of QTLs for high grain Fe and Zn, yield and yield components helps in precise and faster development of high Fe and Zn rice. We carried out a three-season evaluation using IR05F102 x IR69428 derived doubled-haploid population at IRRI. Inclusive composite interval mapping was carried out using SNP markers and Best Linear Unbiased Estimates of the phenotypic traits. A total of 23 QTLs were identified for eight agronomic traits and grain Fe and Zn concentration that explained 7.2 to 22.0% PV. A QTL by environment interaction analysis confirmed the stability of nine QTLs, including two QTLs for Zn on chromosomes 5 and 12. One epistatic interaction for plant height was significant with 28.4% PVE. Moreover, five QTLs were identified for Fe and Zn that harbor several candidate genes, e.g. OsZIP6 on QTL qZn5.1. A number of QTLs were associated with a combination of greater yield and increased grain Zn levels. These results are useful for development of new rice varieties with good agronomic traits and high grain Zn using MAS, and identification of genetic resources with the novel QTLs for grain Zn.

Zinc and Iron Nutrition Status in the Philippines Population and Local Soils.

The Philippines is one of the major rice-producing and rice-consuming countries of Asia. A large portion of its population depends on rice for their daily caloric intake and nutritional needs. The lack of dietary diversity among poor communities has led to nutritional consequences, particularly micronutrient deficiencies. Iron-deficiency anemia (IDA) and zinc deficiency (ZnD) are two serious nutritional problems that affect the health and economic sector of the country. Since rice dominates the Filipino diet by default, biofortification of rice will help improve the micronutrient status. The Philippine government has proactively initiated various programs and policies to address micronutrient deficiencies, particularly through fortification of basic food commodities. Biofortification, the fortification of rice with micronutrients through breeding, is considered the most sustainable and cost-effective strategy that can benefit large vulnerable populations. However, developing promising genotypes with micronutrient-enriched grains should be coupled with improving micronutrient bioavailability in the soil in order to optimize biofortification. This review documents the prevailing soil Zn-deficiency problems in the major rice production areas in the Philippines that may influence the Zn nutritional status of the population. The article also reports on the biofortification efforts that have resulted in the development of two biofortified varieties approved for commercial release in the Philippines. As nutritional security is increasingly recognized as a priority area, greater efforts are required to develop biofortified rice varieties that suit both farmers' and consumers' preferences, and that can address these critical needs for human health in a sustainable and cost-effective manner.

Genetic Dissection of Grain Nutritional Traits and Leaf Blight Resistance in Rice.

Colored rice is rich in nutrition and also a good source of valuable genes/quantitative trait loci (QTL) for nutrition, grain quality, and pest and disease resistance traits for use in rice breeding. Genome-wide association analysis using high-density single nucleotide polymorphism (SNP) is useful in precisely detecting QTLs and genes. We carried out genome-wide association analysis in 152 colored rice accessions, using 22,112 SNPs to map QTLs for nutritional, agronomic, and bacterial leaf blight (BLB) resistance traits. Wide variations and normal frequency distributions were observed for most of the traits except anthocyanin content and BLB resistance. The structural and principal component analysis revealed two subgroups. The linkage disequilibrium (LD) analysis showed 74.3% of the marker pairs in complete LD, with an average LD distance of 1000 kb and, interestingly, 36% of the LD pairs were less than 5 Kb, indicating high recombination in the panel. In total, 57 QTLs were identified for ten traits at p < 0.0001, and the phenotypic variance explained (PVE) by these QTLs varied from 9% to 18%. Interestingly, 30 (53%) QTLs were co-located with known or functionally-related genes. Some of the important candidate genes for grain Zinc (Zn) and BLB resistance were OsHMA9, OsMAPK6, OsNRAMP7, OsMADS13, and OsZFP252, and Xa1, Xa3, xa5, xa13 and xa26, respectively. Red rice genotype, Sayllebon, which is high in both Zn and anthocyanin content, could be a valuable material for a breeding program for nutritious rice. Overall, the QTLs identified in our study can be used for QTL pyramiding as well as genomic selection. Some of the novel QTLs can be further validated by fine mapping and functional characterization. The results show that pigmented rice is a valuable resource for mineral elements and antioxidant compounds; it can also provide novel alleles for disease resistance as well as for yield component traits. Therefore, large opportunities exist to further explore and exploit more colored rice accessions for use in breeding.

Genome-Wide Association Mapping in a Rice MAGIC Plus Population Detects QTLs and Genes Useful for Biofortification.

The development of rice genotypes with micronutrient-dense grains and disease resistance is one of the major priorities in rice improvement programs. We conducted Genome-wide association studies (GWAS) using a Multi-parent Advanced Generation Inter-Cross (MAGIC) Plus population to identify QTLs and SNP markers that could potentially be integrated in biofortification and disease resistance breeding. We evaluated 144 MAGIC Plus lines for agronomic and biofortification traits over two locations for two seasons, while disease resistance was screened for one season in the screen house. X-ray fluorescence technology was used to measure grain Fe and Zn concentrations. Genotyping was carried out by genotype by sequencing and a total of 14,242 SNP markers were used in the association analysis. We used Mixed linear model (MLM) with kinship and detected 57 significant genomic regions with a -log10 (P-value) ≥ 3.0. The PH 1.1 and Zn 7.1 were consistently identified in all the four environments, ten QTLs qDF 3.1, qDF 6.2 qDF 9.1 qPH 5.1 qGL 3.1, qGW 3.1, qGW 11.1, and qZn 6.2 were detected in two environments, while two major loci qBLB 11.1 and qBLB 5.1 were identified for Bacterial Leaf Blight (BLB) resistance. The associated SNP markers were found to co-locate with known major genes and QTLs such as OsMADS50 for days to flowering, osGA20ox2 for plant height, and GS3 for grain length. Similarly, Xa4 and xa5 genes were identified for BLB resistance and Pi5(t), Pi28(t), and Pi30(t) genes were identified for Blast resistance. A number of metal homeostasis genes OsMTP6, OsNAS3, OsMT2D, OsVIT1, and OsNRAMP7 were co-located with QTLs for Fe and Zn. The marker-trait relationships from Bayesian network analysis showed consistency with the results of GWAS. A number of promising candidate genes reported in our study can be further validated. We identified several QTLs/genes pyramided lines with high grain Zn and acceptable yield potential, which are a good resource for further evaluation to release as varieties as well as for use in breeding programs.

Identification of genomic regions associated with agronomic and biofortification traits in DH populations of rice.

Rice provides energy and nutrition to more than half of the world's population. Breeding rice varieties with the increased levels of bioavailable micronutrients is one of the most sustainable approaches to tackle micronutrient malnutrition. So, high zinc and iron content in the grain are primary targets in rice biofortification breeding. In this study, we conducted QTL mapping using doubled haploid (DH) populations, PSBRc82 x Joryeongbyeo and PSBRc82 x IR69428, phenotyped for agronomic traits and micronutrients during two growing seasons and using genotypic information from analysis with the 6K SNP chip. A number of DH lines were identified as having high grain Zn and Fe content in polished rice. Importantly, we identified 20 QTLs for agronomic traits and 59 QTLs for a number of biofortification traits. Of the 79 QTLs, 12 were large-effect QTLs (>25% PVE), nine QTLs were consistent across seasons in either population, and one QTL was identified in both populations. Moreover, at least two QTLs were clustered in defined regions of chromosomes 1, 2, 3, 4, 5, 7 and 9. Eight epistatic interactions were detected for Cu, Mg, Na, and Zn in population 1. Furthermore, we identified several candidate genes near QTLs for grain Zn (OsNRAMP, OsNAS, OsZIP, OsYSL, OsFER, and OsZIFL family) and grain yield (OsSPL14 and OsSPL16). These new QTLs and candidate genes help to further elucidate the genetic basis for grain micronutrient concentration, and may prove useful for marker assisted breeding for this important trait.

Advances in breeding for high grain Zinc in Rice.

Zinc (Zn) is one of the most essential micronutrients required for the growth and development of human beings. More than one billion people, particularly children and pregnant women suffer from Zn deficiency related health problems in Asia. Rice is the major staple food for Asians, but the presently grown popular high yielding rice varieties are poor supplier of Zn in their polished form. Breeding rice varieties with high grain Zn has been suggested to be a sustainable, targeted, food-based and cost effective approach in alleviating Zn deficiency. The physiological, genetic and molecular mechanisms of Zn homeostasis have been well studied, but these mechanisms need to be characterized from a biofortification perspective and should be well integrated with the breeding processes. There is a significant variation for grain Zn in rice germplasm and efforts are being directed at exploiting this variation through breeding to develop high Zn rice varieties. Several QTLs and gene specific markers have been identified for grain Zn and there is a great potential to use them in Marker-Assisted Breeding. A thorough characterization of genotype and environmental interactions is essential to identify key environmental factors influencing grain Zn. Agronomic biofortification has shown inconsistent results, but a combination of genetic and agronomic biofortification strategies may be more effective. Significant progress has been made in developing high Zn rice lines for release in target countries. A holistic breeding approach involving high Zn trait development, high Zn product development, product testing and release, including bioefficacy and bioavailability studies is essential for successful Zn biofortification.