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.

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 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.

Methodology: ssb-MASS: a single seed-based sampling strategy for marker-assisted selection in rice.

BACKGROUND: Integrated breeding approaches such as combining marker-assisted selection and rapid line fixation through single-seed-descent, can effectively increase the frequency of desirable alleles in a breeding program and increase the rate of genetic gain for quantitative traits by shortening the breeding cycle. However, with most genotyping being outsourced to 3rd party service providers' nowadays, sampling has become the bottleneck for many breeding programs. While seed-chipping as prevailed as an automatable seed sampling protocol in many species, the symmetry of rice seeds makes this solution as laborious and costly as sampling leaf tissue. The aim of this study is to develop, validate and deploy a single seed sampling strategy for marker-assisted selection of fixed lines in rice that is more efficient, cost-effective and convenient compared to leaf-based sampling protocols without compromising the accuracy of the marker-assisted selection results. RESULTS: Evaluations replicated across accessions and markers showed that a single rice seed is sufficient to generate enough DNA (7-8 ng/µL) to run at least ten PCR trait-markers suitable for marker-assisted selection strategies in rice. The DNA quantity and quality extracted from single seeds from fixed lines (F6) with different physical and/or chemical properties were not significantly different. Nor were there significant differences between single seeds collected 15 days after panicle initiation compared to those harvested at maturity. A large-scale comparison between single seed and leaf-based methodologies showed not only high levels of genotypic concordance between both protocols (~ 99%) but also higher SNP call rates in single seed (99.24% vs. 97.5% in leaf). A cost-benefit analysis showed that this single seed sampling strategy decreased the cost of sampling fourfold. An advantage of this approach is that desirable genotypes can be selected before investing in planting activities reducing the cost associated with field operations. CONCLUSION: This study reports the development of a cost-effective and simple single seed genotyping strategy that facilitates the adoption and deployment of marker-assisted selection strategies in rice. This will allow breeders to increase the frequency of favorable alleles and combine rapid generation advancement techniques much more cost-effectively accelerating the process and efficiency of parental selection and varietal development.

Decision tools for bacterial blight resistance gene deployment in rice-based agricultural ecosystems.

Attempting to achieve long-lasting and stable resistance using uniformly deployed rice varieties is not a sustainable approach. The real situation appears to be much more complex and dynamic, one in which pathogens quickly adapt to resistant varieties. To prevent disease epidemics, deployment should be customized and this decision will require interdisciplinary actions. This perspective article aims to highlight the current progress on disease resistance deployment to control bacterial blight in rice. Although the model system rice-Xanthomonas oryzae pv. oryzae has distinctive features that underpin the need for a case-by-case analysis, strategies to integrate those elements into a unique decision tool could be easily extended to other crops.