SpeedFlower: a comprehensive speed breeding protocol for indica and japonica rice.

To increase rice yields and feed billions of people, it is essential to enhance genetic gains. However, the development of new varieties is hindered by longer generation times and seasonal constraints. To address these limitations, a speed breeding facility has been established and a robust speed breeding protocol, SpeedFlower is developed that allows growing 4-5 generations of indica and/or japonica rice in a year. Our findings reveal that a high red-to-blue (2R > 1B) spectrum ratio, followed by green, yellow and far-red (FR) light, along with a 24-h long day (LD) photoperiod for the initial 15 days of the vegetative phase, facilitated early flowering. This is further enhanced by 10-h short day (SD) photoperiod in the later stage and day and night temperatures of 32/30 °C, along with 65% humidity facilitated early flowering ranging from 52 to 60 days at high light intensity (800 µmol m-2 s-1). Additionally, the use of prematurely harvested seeds and gibberellic acid treatment reduced the maturity duration by 50%. Further, SpeedFlower was validated on a diverse subset of 198 rice accessions from 3K RGP panel encompassing all 12 distinct groups of Oryza sativa L. classes. Our results confirmed that using SpeedFlower one generation can be achieved within 58-71 days resulting in 5.1-6.3 generations per year across the 12 sub-groups. This breakthrough enables us to enhance genetic gain, which could feed half of the world's population dependent on rice.

Molecular and Morphological Characterization of Introgression Lines with Resistance to Bacterial Leaf Blight and Blast in Rice.

The present study evaluates marker assisted forward breeding (MAFB)-derived disease resistant introgression lines (ILs) which do not have the targeted resistance genes for bacterial blight (xa5 + xa13 + Xa21) and blast (Pi2 + Pi9 + Pi54). The ILs were derived in the background of two elite rice cultivars, Krishna Hamsa [Recurrent Parent 1 (RP1)] and WGL 14 (RP2), involving multi-parent inter-crossing. Molecular characterization with gene specific markers for seven reported resistance genes each for bacterial blight (Xa33, Xa38, xa23, Xa4, xa8, Xa27 and Xa41) and blast (Pi1, Pi20, Pi38, Pib, Pitp, Pizt and Pi40) revealed the presence of xa8 and Xa38, in addition to the targeted xa5, xa13 and Xa21 for bacterial blight resistance and Pi1, Pi38, Pi40, Pi20, Pib and Pipt, in addition to the targeted Pi9 and Pi54, for blast resistance in various combinations. A maximum of nine resistance genes xa5 + Xa21 + Pi54 + xa8 + Pipt + Pi38 + Pi1 + Pi20 + Pib was observed in RP1-IL 19030 followed by eight genes xa5 + xa13 + Xa21 + xa8 + Pi9 + Pipt + Pi1 + Pi20 in two RP2-ILs, 19344 and 19347. ANOVA revealed the presence of significant variability for all the yield traits except "days to 50% flowering" (DFF). Box plots depicted the seasonal differences in the phenotypic expression of the yield traits. There was significant positive association of grain yield with days to flowering, tiller number and panicle number. Thousand grain weight is also significantly and positively correlated with grain yield. On the contrary, grain yield showed a significantly negative association with plant height. Multi-parent selective inter-crossing in the present study not only led to the development of high yielding disease resistant ILs but also enhanced recovery of the recurrent parent via selection for essential morphological features. More than 90.0% genetic similarity in the ILs based on SNP-based background selection demonstrated the success of multi-parent selective intercrossing in the development of disease resistant NILs.

Genome-wide association studies for a comprehensive understanding of the genetic architecture of culm strength and yield traits in rice.

Lodging resistance in rice is a complex trait determined by culm morphological and culm physical strength traits, and these traits are a major determinant of yield. We made a detailed analysis of various component traits with the aim of deriving optimized parameters for measuring culm strength. Genotyping by sequencing (GBS)-based genome-wide association study (GWAS) was employed among 181 genotypes for dissecting the genetic control of culm strength traits. The VanRaden kinship algorithm using 6,822 filtered single-nucleotide polymorphisms (SNPs) revealed the presence of two sub-groups within the association panel with kinship values concentrated at<0.5 level, indicating greater diversity among the genotypes. A wide range of phenotypic variation and high heritability for culm strength and yield traits were observed over two seasons, as reflected in best linear unbiased prediction (BLUP) estimates. The multi-locus model for GWAS resulted in the identification of 15 highly significant associations (p< 0.0001) for culm strength traits. Two novel major effect marker-trait associations (MTAs) for section modulus and bending stress were identified on chromosomes 2 and 12 with a phenotypic variance of 21.87% and 10.14%, respectively. Other MTAs were also noted in the vicinity of previously reported putative candidate genes for lodging resistance, providing an opportunity for further research on the biochemical basis of culm strength. The quantitative trait locus (QTL) hotspot identified on chromosome 12 with the synergistic association for culm strength trait (section modulus, bending stress, and internode breaking weight) and grain number can be considered a novel genomic region that can serve a dual purpose of enhancing culm strength and grain yield. Elite donors in the indica background with beneficial alleles of the identified major QTLs could be a valuable resource with greater significance in practical plant breeding programs focusing on improving lodging resistance in rice.

Multiparent-Derived, Marker-Assisted Introgression Lines of the Elite Indian Rice Cultivar, 'Krishna Hamsa' Show Resistance against Bacterial Blight and Blast and Tolerance to Drought.

Major biotic stresses viz., bacterial blight (BB) and blast and brown plant hopper (BPH) coupled with abiotic stresses like drought stress, significantly affect rice yields. To address this, marker-assisted intercross (IC) breeding involving multiple donors was used to combine three BB resistance genes-xa5, xa13 and Xa21, two blast resistance genes-Pi9 and Pi54, two BPH resistance genes-Bph20 and Bph21, and four drought tolerant quantitative trait loci (QTL)-qDTY1.1, qDTY2.1, qDTY3.1 and qDTY12.1-in the genetic background of the elite Indian rice cultivar 'Krishna Hamsa'. Three cycles of selective intercrossing followed by selfing coupled with foreground selection and phenotyping for the target traits resulted in the development of 196 introgression lines (ILs) with a myriad of gene/QTL combinations. Based on the phenotypic reaction, the ILs were classified into seven phenotypic classes of resistance/tolerance to the following: (1) BB, blast and drought-5 ILs; (2) BB and blast-10 ILs; (3) BB and drought-9 ILs; (4) blast and drought-42 ILs; (5) BB-3 ILs; (6) blast-84 ILs; and (7) drought-43 ILs; none of the ILs were resistant to BPH. Positive phenotypic response (resistance) was observed to both BB and blast in 2 ILs, BB in 9 ILs and blast in 64 ILs despite the absence of corresponding R genes. Inheritance of resistance to BB and/or blast in such ILs could be due to the unknown genes from other parents used in the breeding scheme. Negative phenotypic response (susceptibility) was observed in 67 ILs possessing BB-R genes, 9 ILs with blast-R genes and 9 ILs harboring QTLs for drought tolerance. Complex genic interactions and recombination events due to the involvement of multiple donors explain susceptibility in some of the marker positive ILs. The present investigation successfully demonstrates the possibility of rapid development of multiple stress-tolerant/resistant ILs in the elite cultivar background involving multiple donors through selective intercrossing and stringent phenotyping. The 196 ILs in seven phenotypic classes with myriad of gene/QTL combinations will serve as a useful genetic resource in combining multiple biotic and abiotic stress resistance in future breeding programs.

Marker-assisted forward breeding to develop a drought-, bacterial-leaf-blight-, and blast-resistant rice cultivar.

Among the different challenges related to rice (Oryza sativa L.) cultivation, drought, bacterial leaf blight (BLB), and blast are the key stresses that significantly affect grain yield (GY) in rice. To ameliorate this issue, marker-assisted forward breeding (MAFB) coupled with a simultaneous crossing approach was used to combine three drought tolerant quantitative trait loci (QTL)-qDTY1.1 , qDTY3.1 , and qDTY12.1 -four BLB genes-Xa4, xa5, xa13, and Xa21-and one blast-resistance gene, Pi9, in the elite rice cultivar Lalat. The introgression lines (ILs) developed in the current study were phenotypically screened for drought, BLB, and blast resistance at the F7 -F8 generation. Under the reproductive stage (RS) drought stress, the yield advantage of ILs, with major-effect QTL (qDTY) over elite parent Lalat, ranges from 9 to 124% in DS2019 and from 7 to 175% in WS2019. The selected ILs were highly resistant to BLB, with lesion lengths ranging from 1.3 to 3.0 cm and blast scores ranging from 1 to 3. ILs that were tolerant to RS drought, resistant to BLB, and blast disease and had similar or higher yields than Lalat were analyzed for grain quality. Six ILs were found to have similar grain quality characteristics to Lalat including hulling, milling, head rice recovery (HRR), chalkiness, alkali spreading value (ASV), and amylose content (AC). This study showed that MAFB, together with simultaneous crossing, would be an effective strategy to rapidly combine multiple stresses in rice. The ILs developed in this study could help to ensure yield sustainability in rainfed environments or be used as genetic material in future breeding programs.

Marker-assisted forward and backcross breeding for improvement of elite Indian rice variety Naveen for multiple biotic and abiotic stress tolerance.

The elite Indian rice variety, Naveen is highly susceptible to major biotic and abiotic stresses such as blast, bacterial blight (BB), gall midge (GM) and drought which limit its productivity in rainfed areas. In the present study, a combined approach of marker-assisted forward (MAFB) and back cross (MABC) breeding was followed to introgress three major genes, viz., Pi9 for blast, Xa21 for bacterial blight (BB), and Gm8 for gall midge (GM) and three major QTLs, viz., qDTY1.1, qDTY2.2 and qDTY4.1 conferring increased yield under drought in the background of Naveen. At each stage of advancement, gene-based/linked markers were used for the foreground selection of biotic and abiotic stress tolerant genes/QTLs. Intensive phenotype-based selections were performed in the field for identification of lines with high level of resistance against blast, BB, GM and drought tolerance without yield penalty under non-stress situation. A set of 8 MAFB lines and 12 MABC lines with 3 to 6 genes/QTLs and possessing resistance/tolerance against biotic stresses and reproductive stage drought stress with better yield performance compared to Naveen were developed. Lines developed through combined MAFB and MABC performed better than lines developed only through MAFB. This study exemplifies the utility of the combined approach of marker-assisted forward and backcrosses breeding for targeted improvement of multiple biotic and abiotic stress resistance in the background of popular mega varieties.

Rice breeding for yield under drought has selected for longer flag leaves and lower stomatal density.

Direct selection for yield under drought has resulted in the release of a number of drought-tolerant rice varieties across Asia. In this study, we characterized the physiological traits that have been affected by this strategy in breeding trials across sites in Bangladesh, India, and Nepal. Drought- breeding lines and drought-tolerant varieties showed consistently longer flag leaves and lower stomatal density than our drought-susceptible check variety, IR64. The influence of environmental parameters other than drought treatments on leaf traits was evidenced by close grouping of treatments within a site. Flag-leaf length and width appeared to be regulated by different environmental parameters. In separate trials in the Philippines, the same breeding lines studied in South Asia showed that canopy temperature under drought and harvest index across treatments were most correlated with grain yield. Both atmospheric and soil stress strengthened the relationships between leaf traits and yield. The stable expression of leaf traits among genotypes and the identification of the environmental conditions in which they contribute to yield, as well as the observation that some breeding lines showed longer time to flowering and higher canopy temperature than IR64, suggest that selection for additional physiological traits may result in further improvements of this breeding pool.

Genetic gain for rice yield in rainfed environments in India.

The complexity of genotype × environment interactions under drought reduces heritability, which determines the effectiveness of selection for drought tolerance and development of drought tolerant varieties. Genetic progress measured through changes in yield performance over time is important in determining the efficiency of breeding programmes in which test cultivars are replaced each year on the assumption that the new cultivars will surpass the older cultivars. The goal of our study was to determine the annual rate of genetic gain for rice grain yield in a drought-prone rainfed system in a series of multi-environment trials conducted from 2005 to 2014 under the Drought Breeding Network of Indian sites in collaboration with the International Rice Research Institute (IRRI). Our results show a positive trend in grain yield with an annual genetic yield increase of about 0.68 % under irrigated control, 0.87 % under moderate reproductive stage drought stress and 1.9 % under severe reproductive stage drought stress due to breeding efforts. The study also demonstrates the effectiveness of direct selection for grain yield under both irrigated control as well as managed drought stress screening to improve yield in typical rainfed systems. IRRI's drought breeding programme has exhibited a significant positive trend in genetic gain for grain yield over the years under both drought stress as well as favorable irrigated control conditions. Several drought tolerant varieties released from the programme have outperformed the currently grown varieties under varied conditions in the rainfed environments on farmers' fields.

Marker Assisted Forward Breeding to Combine Multiple Biotic-Abiotic Stress Resistance/Tolerance in Rice.

BACKGROUND: Unfavorable climatic changes have led to an increased threat of several biotic and abiotic stresses over the past few years. Looking at the massive damage caused by these stresses, we undertook a study to develop high yielding climate-resilient rice, using genes conferring resistance against blast (Pi9), bacterial leaf blight (BLB) (Xa4, xa5, xa13, Xa21), brown planthopper (BPH) (Bph3, Bph17), gall midge (GM) (Gm4, Gm8) and QTLs for drought tolerance (qDTY1.1 and qDTY3.1) through marker-assisted forward breeding (MAFB) approach. RESULT: Seven introgression lines (ILs) possessing a combination of seven to ten genes/QTLs for different biotic and abiotic stresses have been developed using marker-assisted selection (MAS) breeding method in the background of Swarna with drought QTLs. These ILs were superior to the respective recurrent parent in agronomic performance and also possess preferred grain quality with intermediate to high amylose content (AC) (23-26%). Out of these, three ILs viz., IL1 (Pi9+ Xa4+ xa5+ Xa21+ Bph17+ Gm8+ qDTY1.1+ qDTY3.1), IL6 (Pi9+ Xa4+ xa5+ Xa21+ Bph3+ Bph17+ Gm4+ Gm8+ qDTY1.1+ qDTY3.1) and IL7 (Pi9+ Xa4+ xa5+ Bph3+ Gm4+ qDTY1.1+ qDTY3.1) had shown resistance\tolerance for multiple biotic and abiotic stresses both in the field and glasshouse conditions. Overall, the ILs were high yielding under various stresses and importantly they also performed well in non-stress conditions without any yield penalty. CONCLUSION: The current study clearly illustrated the success of MAS in combining tolerance to multiple biotic and abiotic stresses while maintaining higher yield potential and preferred grain quality. Developed ILs with seven to ten genes in the current study showed superiority to recurrent parent Swarna+drought for multiple-biotic stresses (blast, BLB, BPH and GM) together with yield advantages of 1.0 t ha- 1 under drought condition, without adverse effect on grain quality traits under non-stress.

Uncovering of natural allelic variants of key yield contributing genes by targeted resequencing in rice (Oryza sativa L.).

In rice (Oryza sativa L.), during the course of domestication, numerous beneficial alleles remain untapped in the progenitor wild species and landraces. This study aims at uncovering these promising alleles of six key genes influencing the yield, such as DEP1, Ghd7, Gn1a, GS3, qSW5 and sd1 by targeted resequencing of the 200 rice genotypes. In all, 543 nucleotide variations including single nucleotide polymorphisms and insertion and deletion polymorphisms were identified from the targeted genes. Of them, 225 were novel alleles, which identified in the present study only and 91 were beneficial alleles that showed significant association with the yield traits. Besides, we uncovered 128 population-specific alleles with indica being the highest of 79 alleles. The neutrality tests revealed that pleiotropic gene, Ghd7 and major grain size contributing gene, GS3 showed positive and balanced selection, respectively during the domestication. Further, the haplotype analysis revealed that some of the rice genotypes found to have rare haplotypes, especially the high yielding variety, BPT1768 has showed maximum of three genes such as Gn1a-8, qSW5-12 and GS3-29. The rice varieties with novel and beneficial alleles along with the rare haplotypes identified in the present study could be of immense value for yield improvement in the rice breeding programs.

Marker Assisted Breeding to Develop Multiple Stress Tolerant Varieties for Flood and Drought Prone Areas.

BACKGROUND: Climate extremes such as drought and flood have become major constraints to the sustainable rice crop productivity in rainfed environments. Availability of suitable climate-resilient varieties could help farmers to reduce the grain yield losses resulting from the climatic extremities. The present study was undertaken with an aim to develop high-yielding drought and submergence tolerant rice varieties using marker assisted introgression of qDTY1.1, qDTY2.1, qDTY3.1 and Sub1. Performance of near isogenic lines (NILs) developed in the background of Swarna was evaluated across 60 multi-locations trials (MLTs). The selected promising lines from MLTs were nominated and evaluated in national trials across 18 locations in India and 6 locations in Nepal. RESULTS: Grain yield advantage of the NILs with qDTY1.1 + qDTY2.1 + qDTY3.1 + Sub1 and qDTY2.1 + qDTY3.1 + Sub1 ranged from 76 to 2479 kg ha- 1 and 396 to 2376 kg ha- 1 under non-stress (NS) respectively and 292 to 1118 kg ha- 1 and 284 to 2086 kg ha- 1 under reproductive drought stress (RS), respectively. The NIL, IR96322-34-223-B-1-1-1-1 having qDTY1.1 + qDTY2.1 + qDTY3.1 + Sub1 has been released as variety CR dhan 801 in India. IR 96321-1447-651-B-1-1-2 having qDTY1.1 + qDTY3.1 + Sub 1 and IR 94391-131-358-19-B-1-1-1 having qDTY3.1 + Sub1 have been released as varieties Bahuguni dhan-1' and 'Bahuguni dhan-2' respectively in Nepal. Background recovery of 94%, 93% and 98% was observed for IR 96322-34-223-B-1-1-1-1, IR 96321-1447-651-B-1-1-2 and IR 94391-131-358-19-B-1-1-1 respectively on 6 K SNP Infinium chip. CONCLUSION: The drought and submergence tolerant rice varieties with pyramided multiple QTLs can ensure 0.2 to 1.7 t ha- 1 under reproductive stage drought stress and 0.1 to 1.0 t ha- 1 under submergence conditions with no yield penalty under non-stress to farmers irrespective of occurrence of drought and/or flood in the same or different seasons.

Incorporation of the novel bacterial blight resistance gene Xa38 into the genetic background of elite rice variety Improved Samba Mahsuri.

Bacterial blight (BB) in rice caused by Xanthomonas oryzae pv. oryzae (Xoo) is a major global production constraint, particularly in irrigated and rain-fed lowland areas. Improved Samba Mahsuri (ISM) is an elite, high-yielding, fine-grain type, BB-resistant rice variety possessing three BB-resistant genes (Xa21, xa13 and xa5) and is highly popular in the southern parts of India. As the BB pathogen is highly dynamic and the evolution of pathogen virulence against the deployed resistance genes is common, we added a novel BB-resistant gene, Xa38, into ISM through marker-assisted backcross breeding (MABB) to increase the spectrum and durability of BB resistance. The breeding line PR 114 (Xa38) was used as the donor for Xa38, whereas ISM was used as the recurrent parent. Foreground selection was conducted using PCR-based gene-specific markers for the target genes, whereas background selection was conducted using a set of polymorphic SSR markers between the parents and backcrossing that continued until the third generation. Eighteen homozygous BC3F2 plants possessing all four BB-resistant genes in the homozygous state and with a recurrent parent genome (RPG) recovery of more than 92% were identified and advanced to the BC3F6 generation. These 18 backcross-derived lines (BDLs) exhibited very high level of resistance against multiple Xoo strains and displayed agro-morphological traits, grain qualities and yield levels similar to or better than those of the recurrent parent ISM.

Genotype × Environment Interactions of Yield Traits in Backcross Introgression Lines Derived from Oryza sativa cv. Swarna/Oryza nivara.

Advanced backcross introgression lines (BILs) developed from crosses of Oryza sativa var. Swarna/O. nivara accessions were grown and evaluated for yield and related traits. Trials were conducted for consecutive three seasons in field conditions in a randomized complete block design with three replications. Data on yield traits under irrigated conditions were analyzed using the Additive Main Effect and Multiplicative Interaction (AMMI), Genotype and Genotype × Environment Interaction (GGE) and modified rank-sum statistic (YSi) for yield stability. BILs viz., G3 (14S) and G6 (166S) showed yield stability across the seasons along with high mean yield performance. G3 is early in flowering with high yield and has good grain quality and medium height, hence could be recommended for most of the irrigated locations. G6 is a late duration genotype, with strong culm strength, high grain number and panicle weight. G6 has higher yield and stability than Swarna but has Swarna grain type. Among the varieties tested DRRDhan 40 and recurrent parent Swarna showed stability for yield traits across the seasons. The component traits thousand grain weight, panicle weight, panicle length, grain number and plant height explained highest genotypic percentage over environment and interaction factors and can be prioritized to dissect stable QTLs/ genes. These lines were genotyped using microsatellite markers covering the entire rice genome and also using a set of markers linked to previously reported yield QTLs. It was observed that wild derived lines with more than 70% of recurrent parent genome were stable and showed enhanced yield levels compared to genotypes with higher donor genome introgressions.

Genetic improvement of biofuel plants: recent progress and patents.

Due to depleting reserves of fossil fuels, political uncertainties, increase in demand of energy needs and growing concerns of environmental effects, bioenergy as an alternative source of energy needs had taken centre stage globally. In this report, we review the progress made in lignocellulose, cellulose and fermentation based biofuels in addition to tree borne oil seeds. Algae as a source of feedstock for the biofuel has also been reviewed. Recent efforts in genome sequencing of biofuel crops and molecular breeding approaches have increased our understanding towards crop improvement of major feedstocks. Besides, patenting trends in bioenergy sector were assessed by patent landscape analysis. The results showed an increasing trend in published patents during the last decade which is maximum during 2011. A conceptual framework of "transgenesis in biofuels to industrial application" was developed based on the patent analytics viz., International Patent Classification (IPC) analysis and Theme Maps. A detailed claim analysis based on the conceptual framework assessed the patenting trends that provided an exhaustive dimension of the technology. The study emphasizes the current thrust in bioenergy sector by various public and private institutions to expedite the process of biofuel production.