Mapping QTLs for yield and photosynthesis-related traits in three consecutive backcross populations of Oryza sativa cultivar Cottondora Sannalu (MTU1010) and Oryza rufipogon.

MAIN CONCLUSION: Identification of trait enhancing QTLs for yield and photosynthesis-related traits in rice using interspecific mapping population and chromosome segment substitution lines derived from a cross between Oryza sativa and Oryza rufipogon. Wild rice contains novel genes which can help in improving rice yield. Common wild rice Oryza rufipogon is a known source for enhanced photosynthesis and yield-related traits. We developed BC2F2:3:4 mapping populations using O. rufipogon IC309814 with high photosynthetic rate as donor, and elite cultivar MTU1010 as recurrent parent. Evaluation of 238 BC2F2 families for 13 yield-related traits and 208 BC2F2 families for seven photosynthesis-related physiological traits resulted in identification of significantly different lines which performed better than MTU1010 for various yield contributing traits. 49 QTLs were identified for 13 yield traits and 7 QTLs for photosynthesis-related traits in BC2F2. In addition, 34 QTLs in BC2F3 and 26 QTLs in BC2F4 were also detected for yield traits.11 common QTLs were identified in three consecutive generations and their trait-increasing alleles were derived from O. rufipogon. Significantly, one major effect common QTL qTGW3.1 for thousand grain weight with average phenotypic variance 8.1% and one novel QTL qBM7.1 for biomass were identified. Photosynthesis-related QTLs qPN9.1, qPN12.1, qPN12.2 qSPAD1.1 and qSPAD6.1 showed additive effect from O. rufipogon. A set of 145 CSSLs were identified in BC2F2 which together represented 87% of O. rufipogon genome. In addition, 87 of the 145 CSSLs were significantly different than MTU1010 for at least one trait. The major effect QTLs can be fine mapped for gene discovery. CSSLs developed in this study are a good source of novel alleles from O. rufipogon in the background of Cottondora Sannalu for rapid improvement of any trait in rice.

Detecting CSSLs and yield QTLs with additive, epistatic and QTL×environment interaction effects from Oryza sativa × O. nivara IRGC81832 cross.

Chromosome segment substitution lines (CSSLs) are useful tools for precise mapping of quantitative trait loci (QTLs) and the evaluation of gene action and interaction in inter-specific crosses. In this study, a set of 90 back cross lines at BC2F8 generation derived from Swarna x Oryza nivara IRGC81832 was evaluated for yield traits under irrigated conditions in wet seasons of 3 consecutive years. We identified a set of 70 chromosome segment substitution lines, using genotyping data from 140 SSR markers covering 94.4% of O. nivara genome. Among these, 23 CSSLs were significantly different for 7 traits. 22 QTLs were detected for 11 traits with 6.51 to 46.77% phenotypic variation in 90 BILs. Three pleiotropic genomic regions associated with yield traits were mapped on chromosomes 1, 8 and 11. The marker interval RM206-RM144 at chromosome 11 was recurrently detected for various yield traits. Ten QTLs were identified consistently in the three consecutive years of testing. Seventeen pairs of significant epistatic QTLs (E-QTLs) were detected for days to flowering, days to maturity and plant height. Chromosome segments from O. nivara contributed trait enhancing alleles. The significantly improved lines and the stable QTLs identified in this study are valuable resource for gene discovery and yield improvement.

Development and use of chromosome segment substitution lines as a genetic resource for crop improvement.

KEY MESSAGE: CSSLs are a complete library of introgression lines with chromosomal segments of usually a distant genotype in an adapted background and are valuable genetic resources for basic and applied research on improvement of complex traits. Chromosome segment substitution lines (CSSLs) are genetic stocks representing the complete genome of any genotype in the background of a cultivar as overlapping segments. Ideally, each CSSL has a single chromosome segment from the donor with a maximum recurrent parent genome recovered in the background. CSSL development program requires population-wide backcross breeding and genome-wide marker-assisted selection followed by selfing. Each line in a CSSL library has a specific marker-defined large donor segment. CSSLs are evaluated for any target phenotype to identify lines significantly different from the parental line. These CSSLs are then used to map quantitative trait loci (QTLs) or causal genes. CSSLs are valuable prebreeding tools for broadening the genetic base of existing cultivars and harnessing the genetic diversity from the wild- and distant-related species. These are resources for genetic map construction, mapping QTLs, genes or gene interactions and their functional analysis for crop improvement. In the last two decades, the utility of CSSLs in identification of novel genomic regions and QTL hot spots influencing a wide range of traits has been well demonstrated in food and commercial crops. This review presents an overview of how CSSLs are developed, their status in major crops and their use in genomic studies and gene discovery.

Characterization of backcross introgression lines derived from Oryza nivara accessions for photosynthesis and yield.

Improvement of photosynthetic traits is a promising strategy to break the yield potential barrier of major food crops. Leaf photosynthetic traits were evaluated in a set of high yielding Oryza sativa, cv. Swarna × Oryza nivara backcross introgression lines (BILs) along with recurrent parent Swarna, both in wet (Kharif) and dry (Rabi) seasons in normal irrigated field conditions. Net photosynthesis (P N) ranged from 15.37 to 23.25 µmol (CO2) m-2 s-1 in the BILs. Significant difference in P N was observed across the seasons and genotypes. Six BILs showed high photosynthesis compared with recurrent parent in both seasons. Chlorophyll content showed minimum variation across the seasons for any specific BIL but significant variation was observed among BILs. Significant positive association between photosynthetic traits and yield traits was observed, but this association was not consistent across seasons mainly due to contrasting weather parameters in both seasons. BILs 166s and 248s with high and consistent photosynthetic rate exhibited stable high yield levels in both the seasons compared to the recurrent parent Swarna. There is scope to exploit photosynthetic efficiency of wild and weedy rice to identify genes for improvement of photosynthetic rate in cultivars.

Identification of Major Effect QTLs for Agronomic Traits and CSSLs in Rice from Swarna/Oryza nivara Derived Backcross Inbred Lines.

Backcross inbred lines (BILs) derived from elite x wild crosses are very useful for basic studies and breeding. The aim of this study was to map quantitative trait loci (QTLs) associated with yield and related traits and to identify chromosomal segment substitution lines (CSSLs) from unselected BC2F8 BILs of Swarna/Oryza nivara IRGC81848. In all, 94 BILs were field evaluated in 2 years (wet seasons, 2014 and 2015) for nine traits; days to 50% flowering, days to maturity (DM), plant height (PH), number of tillers, number of productive tillers, panicle weight, yield per plant, bulk yield, and biomass. BILs were genotyped using 111 polymorphic simple sequence repeats distributed across the genome. Fifteen QTLs including 10 novel QTLs were identified using composite interval mapping, Inclusive composite interval mapping and multiple interval mapping (MIM). O. nivara alleles were trait-enhancing in 26% of QTLs. Only 3 of 15 QTLs were also reported previously in BC2F2 of the same cross. These three included the two major effect QTLs for DM and PH detected in both years with 13 and 20% phenotypic variance. Further, a set of 74 CSSLs was identified using CSSL Finder and 22 of these showed significantly higher values than Swarna for five yield traits. CSSLs, 220S for panicle weight and 10-2S with consistent high yield in both years are worthy of large scale field evaluation. The major QTLs and 22 significantly different CSSLs are a useful resource for rice improvement and dissecting yield related traits.

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 characterization and population structure of Indian rice cultivars and wild genotypes using core set markers.

Genetic diversity among 23 rice genotypes including wild species and cultivars of indica, japonica, aus and aromatic type was investigated using 165 genomewide core set microsatellite (SSR) markers. This genotypic characterization was undertaken to know the genetic similarity among the parental lines to be used in developing a set of chromosome segment substitution lines. In all, 253 alleles were identified using 77 polymorphic SSRs, and polymorphism information content ranged from 0.31 to 0.97 with a mean of 0.79. Cluster analysis grouped the genotypes into three clusters at a genetic similarity of 0.26-0.75. Wild accessions grouped together in cluster-I, indica cultivars formed cluster-II, and aromatic, japonica and aus types came under cluster-III. Principal component analysis also showed similar results. The genotypic data was analyzed using STRUCTURE, and genotypes were grouped into four populations. RM1018 on chromosome 4, RM8009 on chromosome 7, and RM273 on chromosome 12 amplified alleles specific to wild accessions. The information obtained from core set markers would help in selecting diverse parents including wild accessions and for tracking alleles in mapping or breeding populations.

Quantitative real-time PCR assay for rapid detection of plant and human pathogenic Macrophomina phaseolina from field and environmental samples.

A real-time qPCR assay was developed to detect and quantify Macrophomina phaseolina abundance in rhizosphere soil and plant tissue. Both TaqMan and SYBR green techniques were targeted on ~ 1 kb sequence characterized amplified region (SCAR) of M. phaseolina and two sets of specific primers were designed for SYBR green (MpSyK) and TaqMan (MpTqK) assays. No cross-hybridization and no fluorescent signal exceeding the baseline threshold was observed in TaqMan and SYBR green assays, respectively. The minimum detection limit or sensitivity of TaqMan assay was 30 fg/µL of M. phaseolina DNA and limit of quantification of M. phaseolina viable population was estimated as 0.66 × 10(5) CFU/g soil(-1) equivalent to 10 pg/µL of target DNA. This is the first report which demonstrated real-time qPCR assays with greater specificity and sensitivity to detect M. phaseolina population in soil and plant materials.