Genes Xa21 Research Articles

Brown Planthopper, Nilaparvata lugens Stål Resistance in Backcross Derived Rice Lines

A prominent rice pest, brown planthopper (BPH) significantly reduces the grain yield in rice across the globe and employing chemical pesticides leads to unwarranted environmental issues. Breeding for BPH resistance is an essential strategy to mitigate the losses caused by them. Host plant resistance through marker assisted selection is a chief strategy to lessen harms caused by BPH and boost rice production. In this study, we have analyzed BPH resistance in the BC1F5 population, which is a backcross derivative of improved CO51 and Ptb33. Improved CO51 has already been introgressed with bacterial blight resistant genes xa5, xa13 and Xa21 and blast resistant gene Pi54 via marker assisted selection (MAS). Ptb33 was used as the donor parent to incorporate BPH resistant genes bph2 and Bph32 to this CO51 background. The genotypically and phenotypically selected 26 lines of BC1F5 generation were screened against BPH along with parents and checks. The bioassay of the population exhibited a range of variation for BPH resistance. Among the 26 near isogenic lines, 18 (2 resistant and 16 are moderately resistant) and eight showed susceptible to moderate susceptible reaction. The 18 resistant lines were further multiplied and are now in hot spot screening.

High-efficiency prime editing enables new strategies for broad-spectrum resistance to bacterial blight of rice.

Using genetic resistance against bacterial blight (BB) caused by Xanthomonas oryzae pathovar oryzae (Xoo) is a major objective in rice breeding programmes. Prime editing (PE) has the potential to create novel germplasm against Xoo. Here, we use an improved prime-editing system to implement two new strategies for BB resistance. Knock-in of TAL effector binding elements (EBE) derived from the BB susceptible gene SWEET14 into the promoter of a dysfunctional executor R gene xa23 reaches 47.2% with desired edits including biallelic editing at 18% in T0 generation that enables an inducible TALE-dependent BB resistance. Editing the transcription factor TFIIA gene TFIIAγ5 required for TAL effector-dependent BB susceptibility recapitulates the resistance of xa5 at an editing efficiency of 88.5% with biallelic editing rate of 30% in T0 generation. The engineered loci provided resistance against multiple Xoo strains in T1 generation. Whole-genome sequencing detected no OsMLH1dn-associated random mutations and no off-target editing demonstrating high specificity of this PE system. This is the first-ever report to use PE system to engineer resistance against biotic stress and to demonstrate knock-in of 30-nucleotides cis-regulatory element at high efficiency. The new strategies hold promises to fend rice off the evolving Xoo strains and protect it from epidemics.

Marker-Assisted Improvement for Durable Bacterial Blight Resistance in Aromatic Rice Cultivar HUR 917 Popular in Eastern Parts of India.

Bacterial blight (BB) is a devastating disease of rice in the tropics of Indian sub-continent, where the presence of Xoo races with varying levels of genetic diversity and virulence renders disease management extremely challenging. In this context, marker-assisted improvement of plant resistance has been proven as one of the most promising approaches for the development of sustainable rice cultivars. The present study demonstrates the marker-assisted introgression of the three BB resistant genes (Xa21 + xa13 + xa5) into the background of HUR 917, a popular aromatic short grain (ASG) rice cultivar in India. The performance of the resulting improved products (near isogenic lines (NILs), HR 23-5-37-83-5, HR 23-5-37-121-10, HR 23-5-37-121-14, HR 23-65-6-191-13, HR 23-65-6-237-2, HR 23-65-6-258-10 and HR 23-65-6-258-21) establishes the utility of marker-assisted selection (MAS) approach for accelerated trait introgression in rice. The MAS-bred lines carrying three introgressed genes showed broad spectrum BB resistance (lesion length, LL of 1.06 ± 1.35 cm to 4.61 ± 0.87 cm). Besides, these improved lines showed the complete product profile of recurrent parent HUR 917 along with the enhanced level of durable BB resistance. The improved introgression lines with durable BB resistance would contribute to sustainable rice production in India, particularly in the Indo-Gangetic plane that has substantial acreage under HUR 917.

The human inactive X chromosome modulates expression of the active X chromosome

The "inactive" X chromosome (Xi) has been assumed to have little impact, in trans, on the "active" X (Xa). To test this, we quantified Xi and Xa gene expression in individuals with one Xa and zero to three Xis. Our linear modeling revealed modular Xi and Xa transcriptomes and significant Xi-driven expression changes for 38% (162/423) of expressed X chromosome genes. By integrating allele-specific analyses, we found that modulation of Xa transcript levels by Xi contributes to many of these Xi-driven changes (≥121 genes). By incorporating metrics of evolutionary constraint, we identified 10 X chromosome genes most likely to drive sex differences in common disease and sex chromosome aneuploidy syndromes. We conclude that human X chromosomes are regulated both in cis, through Xi-wide transcriptional attenuation, and in trans, through positive or negative modulation of individual Xa genes by Xi. The sum of these cis and trans effects differs widely among genes.

Genetic diversity and pathotype profiling of Xanthomonas oryzae pv. oryzae isolates from diverse rice growing ecosystems of Karnataka state of India

Bacterial leaf blight (BLB) disease of rice caused by Xanthomonas oryzae pv. oryzae (Xoo) is one of the significant constraints for rice production. A study was carried out to analyze the virulence, genetic diversity, and population structure of Xoo isolates collected from different rice ecosystems of Karnataka, India. The taxonomic identity of all 54 isolates was confirmed using a Xoo-specific PCR assay. The virulence assay on a set of twenty-eight near-isogenic differentials identified nine pathotype groups (KPthX-1 to KPthX-9). Among them, KPthX-6 was highly virulent, whereas KPthx-2 was less virulent. Differentials with four and five resistance (R) genes (Xa4+xa5+xa13+Xa21, Xa4+Xa7+xa13+Xa21, and Xa4+xa5+Xa7+xa13+Xa21) were effective against all the pathotypes. Further, genetic diversity was deduced using a universal marker set, JEL-1, and JEL-2, complementary to a conserved repetitive sequence IS1112. The amplicon size and numbers varied from 100 bp to 3 Kb and 1 to 25. Based on the amplicon counts, Jaccard’s co-efficient and phylogenetic analyses were carried out and categorized the 54 isolates into three clusters (I to III). Cluster-II contains the maximum number of isolates (27), followed by cluster-I (23 isolates) and cluster-III (one isolate). The structure analysis categorized the isolates into five subgroups, viz. SG1, SG2, SG3, SG4, and SG5 indicate the existence of five subpopulations. The present study has identified the genetic and virulence diversity of the Xoo population and reported the effective R gene/s for different rice ecosystems, which can be used in the breeding program for the eco-specific management of BLB of rice.

Design and construction of single guide RNA for CRISPR/Cas9 system based on the xa13 resistance gene in some varieties of rice (Oryza sativa)

The xa13 gene is a recessive resistance gene against Xanthomonas oryzae pv. oryzae (Xoo) found in several rice varieties. Activation of this gene will trigger the formation of sucrose as a nutrient supply to Xoo for their growth in the plant. The disruption of this recessive gene expression in the plant can affect the negative impact of the gene, and recently can be created using clustered regularly interspaced short palindromic repeats (CRISPR) system using CRISPR-associated protein-9 (CRISPR/Cas9) technology that requires gRNA to recognize the targeted-sequence. This study aimed to design and construct the gRNA-targeting xa13 gene in rice using bioinformatics tools. CHOPCHOP was used for generated the gRNA candidates according to the target gene sequence. Two candidates of gRNA-targeted xa13 have been selected based on the analysis of bioinformatics data. Each candidate of gRNA consisted of 20 nucleotides (nt) of the target sequence upstream 3 nt of the protospacer adjacent motif (PAM) sequence (5’-NGG) targeting two exons in the xa13 gene. The gRNA1 will target exon 1 and the gRNA2 will target exon 2, with an efficiency of 52.51% and 44.63% respectively. Data showed that the GC content of all gRNA candidates ranged from 55–70% with no target-off location in the whole genome of rice. The transformation and confirmation test based on the physiological and genomic characteristics of transformants confirmed that the design has been successfully constructed.

Molecular Breeding for Incorporation of Submergence Tolerance and Durable Bacterial Blight Resistance into the Popular Rice Variety 'Ranidhan'.

Ranidhan is a popular late-maturing rice variety of Odisha state, India. The farmers of the state suffer heavy loss in years with flash floods as the variety is sensitive to submergence. Bacterial blight (BB) disease is a major yield-limiting factor, and the variety is susceptible to the disease. BB resistance genes Xa21, xa13, and xa5, along with the Sub1 QTL, for submergence stress tolerance were transferred into the variety using marker-assisted backcross breeding approach. Foreground selection using direct and closely linked markers detected the progenies carrying all four target genes in the BC1F1, BC2F1, and BC3F1 generations, and the positive progenies carrying these genes with maximum similarity to the recipient parent, Ranidhan, were backcrossed into each segregating generation. Foreground selection in the BC1F1 generation progenies detected all target genes in 11 progenies. The progeny carrying all target genes and similar to the recipient parent in terms of phenotype was backcrossed, and a total of 321 BC2F1 seeds were produced. Ten progenies carried all target genes/QTL in the BC2F1 generation. Screening of the BC3F1 progenies using markers detected 12 plants carrying the target genes. A total of 1270 BC3F2 seeds were obtained from the best BC3F1 progeny. Foreground selection in the BC3F2 progenies detected four plants carrying the target genes in the homozygous condition. The bioassay of the pyramided lines conferred very high levels of resistance to the predominant isolates of bacterial blight pathogen. These BB pyramided lines were submergence-tolerant and similar to Ranidhan in 13 agro-morphologic and grain quality traits; hence, they are likely to be adopted by farmers.

SCREENING OF RICE LINES OF DIVERSE ORIGIN RESISTANT TO BACTERIAL LEAF BLIGHT UNDER LOCAL CONDITIONS


 
 
 Current study comprised of 20 donor parent lines and four indica elite strains resistant against bacterial leaf blight having diversity in origin viz. Bangladesh, Philippine, Indonesia, Senegal and India. The lines were tested to check resistance against Bacterial Leaf Blight (BLB), having BLB resistance genes Xa-4, xa-5, Xa-7 and xa-13. The experiment was conducted at Agronomic Research Station, Farooqabad, Sheikhupura. Artificial inoculation was done to study disease reaction. DNA marker analysis was also conducted to study the gene status of these lines. Results indicated that seven donor lines were completely resistant to Bacterial Leaf Blight whereas twelve lines exhibited moderate resistance. Among rest of the five susceptible genotypes, three were showing comparatively low susceptibility whereas two completely susceptible to BLB. Most of the lines with three or four gene combinations showing resistant behavior against BLB except AUS-298 that showed moderately susceptible behavior. Combination of genes for Bacterial Leaf Blight resistance can be explored further in targeted research program for disease resistance.
 
 

Molecular marker assisted selection of introgressed four bacterial blight resistance genes (Xa4, Xa7, Xa13 and Xa21) in the genetic background of Mahamaya

Molecular marker assisted selection of introgressed four bacterial blight resistance genes (Xa4, Xa7, Xa13 and Xa21) in the genetic background of Mahamaya

Molecular Marker Assisted Breeding of a New <i>Japonica</i> Hybrid Rice

The development of Japonica  hybrid rice is of great significance to ensure supply of good quality Japonica  rice and food security in China. Aiming at the problems of late growth period, unstable yield, and the need to improve rice quality and seed production yield of Japonica  hybrid rice, a new Japonica  hybrid rice combination 'Shenyou28' with early maturity, good quality, blast resistance, stable and high yield and easy seed production was developed by using molecular breeding techniques including high-density rice gene chip. 'Shenyou28', bred by Shanghai Academy of Agricultural Sciences, is a three-line Japonica  hybrid rice with 'Shen21A' as sterile line and 'Shenhui26-28' as restorer line. For two-year regional trials from 2018 to 2019, the average output were 10 588.5 kg/hm 2 , which was 11.6% higher than the control variety 'Huayou14'. For the one-year production trial, the output reached 10 473.0 kg/hm 2 , which was 4.8% higher than the control variety 'Huayou14'. 'Shenyou28' has good quality, reaching the second grade of national standard, and has fragrance because it carries fragrance gene badh2-p-5 ’ UTR . It also has good resistance to rice blast and bacterial blight because it aggregates the rice blast resistance genes Pi2 , Pita , Pib , Pi9 , Pi54 , Pikm , Pit , and bacterial blight resistance gene Xa21 . It was approved by the Crop Variety Certification Committee of Shanghai in 2020. The breeding process, variety characteristics and key points of high yield mechanized seed production technology of 'Shenyou28' are introduced in this study.

Pyramiding of Four Broad Spectrum Bacterial Blight Resistance Genes in Cross Breeds of Basmati Rice.

Pyramiding of major resistance (R) genes through marker-assisted selection (MAS) is a useful way to attain durable and broad-spectrum resistance against Xanthomonas oryzae pv. oryzae pathogen, the causal agent of bacterial blight (BB) disease in rice (Oryza sativa L.). The present study was designed to pyramid four broad spectrum BB-R genes (Xa4, xa5, xa13 and Xa21) in the background of Basmati-385, an indica rice cultivar with much sought-after qualitative and quantitative grain traits. The cultivar, however, is susceptible to BB and was therefore, crossed with IRBB59 which possesses R genes xa5, xa13 and Xa21, to attain broad and durable resistance. A total of 19 F1 plants were obtained, some of which were backcrossed with Basmati-385 and large number of BC1F1 plants were obtained. In BC1F2 generation, 31 phenotypically superior genotypes having morphological features of Basmati-385, were selected and advanced up to BC1F6 population. Sequence-tagged site (STS)-based MAS was carried out and phenotypic selection was made in each successive generation. In BC1F6 population, potentially homozygous recombinant inbred lines (RILs) from each line were selected and evaluated on the bases of STS evaluation and resistance to local Xanthomonas oryzae pv. oryzae (Xoo) isolates. Line 23 was found pyramided with all four BB-R genes i.e., Xa4, xa5, xa13 and Xa21. Five genotypes including line 8, line 16, line 21, line 27 and line 28 were identified as pyramided with three R genes, Xa4, xa5 and xa13. Pathological study showed that rice lines pyramided with quadruplet or triplet R genes showed the highest level of resistance compared to doublet or singlet R genes. Thus, line 23 with quadruplet, and lines 8, 16, 21, 27, and 28 with triplet R genes, are recommended for replicated yield and resistance trials before release as new rice varieties. Further, traditional breeding coupled with MAS, is a solid way to attain highly effective BB-resistant rice lines with no yield cost.

Pyramiding Submergence Tolerance and Three Bacterial Blight Resistance Genes in Popular Rice Variety Hasanta through Marker-Assisted Backcross Breeding

Flash flooding (FF) along with bacterial blight (BB) outbreak are very destructive for rice production in the rainfed shallow-lowland (RSL) ecosystem. The presence of dynamic Xoo races with varying levels of genetic diversity and virulence renders their management extremely challenging under RSL. In this context, the marker-assisted improvement of plant resistance/tolerance has been proven as one of the most promising strategies towards the development of sustainable cultivars. The present study demonstrates the marker-assisted introgression of the submergence tolerant (Sub1) and three bacterial blight resistant genes (Xa21 + xa13 + xa5) into the genetic background of Hasanta, a long duration popular rice variety in the eastern coastal region of India. The rice genotypes, Swarna Sub1 (carrying Sub1) and IRBB66 (carrying Xa21 + xa13 + Xa7 + xa5 + Xa4) had maximum genetic similarity (0.84 and 0.73, respectively) with Hasanta; recurrent parent (RP) was used as donor. The forward analysis of target genes in F1s, IC1F1s and backcross (BC) generations was performed by linked/genic markers (Sub1bc2; pTA248, xa13prom and RM122), whereas background recovery of RP in each BC and segregating generations was performed utilizing 108 hypervariable SSR markers. Intervened speed breeding (SB) strategy and intensive phenotyping could lead the development of near isogenic lines (NILs) as to the RP in all basic traits. The performance of the near isogenic lines (NILs, BC2F3 and BC2F4), HS 232-411-391-756-37, HS 232-411-391-809-8, HS 232-411-391-756-18, HS 110-224-197-10-36, HS 232-411-391-809-81, HS 110-224-197-10-41 and HS 232-411-391-809-63 establishes the utility of marker-assisted backcross-breeding (MAB) and SB in accelerated trait introgression. The introgressed lines carrying Sub1 + Xa21 + xa13 + xa5 showed 76% to 91% survival under 14 days of submergence and durable BB resistance (percent disease index-PDI of 2.68 ± 0.26 to 6.22 ± 1.08 and lesion length, LL of 1.29 ± 0.12 to 4.2 ± 0.64 cm). Physiological analysis revealed that improved NILs, carrying Sub1 gene conquered adaptive physiological modulations, had reduced the consumption of soluble sugar and the degradation of total chlorophyll contents (TCC), and an enhanced level of Alcohol Dehydrogenase activity (ADH) and proline accumulation in all submergence regimes. The pyramided lines attained complete product profile of RP, that will contribute to sustainable rice production under RSL, particularly in the coastal region that has substantial acreage under the variety Hasanta.

Molecular Breeding of Water-Saving and Drought-Resistant Rice for Blast and Bacterial Blight Resistance.

Rice production is often affected by biotic and abiotic stressors. The breeding of resistant cultivars is a cost-cutting and environmentally friendly strategy to maintain a sustainable high production level. An elite water-saving and drought-resistant rice (WDR), Hanhui3, is susceptible to blast and bacterial blight (BB). This study was conducted to introgress three resistance genes (Pi2, xa5, and Xa23) for blast and BB into Hanhui3, using marker-assisted selection (MAS) for the foreground selection and a whole-genome single-nucleotide polymorphism (SNP) array for the background selection. As revealed by the whole-genome SNP array, the recurrent parent genome (RPG) recovery of the improved NIL was 94.2%. The resistance levels to blast and BB of the improved NIL and its derived hybrids were higher than that of the controls. In addition, the improved NIL and its derived hybrids retained the desired agronomic traits from Hanhui3, such as yield. The improved NIL could be useful to enhance resistance against biotic stressors and produce stable grain yields in Oryza sativa subspecies indica rice breeding programs.

In Vitro Doubled Haploid Production of Bacterial Blight Resistant Plants from BC2F1 Plants (Ranbir Basmati X Pau148) Through Anther Culture

Doubled haploid plants are very important for the development of complete homozygous plants from heterozygous parents in one generation as they possess duplicate copy of haploid chromosome. Haploid production is easily obtained from in vitro anther culture. The present study was undertaken with the objective to develop doubled haploids using anthers for in vitro induction of callus on N6 medium supplemented with various combinations and concentrations of 2,4-dichlorophenoxy acetic acid (2,4-D) (0.5-2.5 mg/L), Kinetin (0.5-1.0 mg/L) and Naphthalene acetic acid (NAA) (2.0 mg/L) as callus induction medium (CIM). The highest callus induction frequency was obtained when N6 medium fortified with 2,4-D (2.5 mg/L), Kinetin (0.5 mg/L) and NAA (2 mg/L) of 10.07 per cent. The induced callus was sub cultured for shoot regeneration on Murashige and Skoog medium (MS) supplemented with growth regulators: Kinetin and NAA (0.5 mg/L each) in combination with BAP (0.0 - 2.5 mg/L). MS medium supplemented with NAA (0.5 mg/L), Kinetin (0.5 mg/L) and BAP (1.5 mg/L) was most responsive exhibiting regeneration frequency of 28.1 per cent which resulted in maximum regeneration of green plantlets and only 5.21 per cent of albinos. Individual plantlets were separated and immersed in liquid MS medium augmented with NAA (0.5-1.0 mg/L) and BAP (0.5-1.0 mg/L). Maximum rooting was observed in MS medium with NAA (0.5 mg/L) and BAP (1.0 mg/L). The survival rate of in-vitro raised plants was 51.51 per cent. Of these surviving plants, 21 plants were observed to have the sterility percentage above 50 percent and hence can be considered as the doubled haploid plants. Plant DH8 is susceptible and DH20 is heterozygous for gene Xa21. Two plants are susceptible for gene xa13

The rice OsERF101 transcription factor regulates the NLR Xa1-mediated immunity induced by perception of TAL effectors.

Plant nucleotide-binding leucine-rich repeat receptors (NLRs) initiate immune responses by recognizing pathogen effectors. The rice gene Xa1 encodes an NLR with an N-terminal BED domain, and recognizes transcription activator-like (TAL) effectors of Xanthomonas oryzae pv oryzae (Xoo). Our goal here was to elucidate the molecular mechanisms controlling the induction of immunity by Xa1. We used yeast two-hybrid assays to screen for host factors that interact with Xa1 and identified the AP2/ERF-type transcription factor OsERF101/OsRAP2.6. Molecular complementation assays were used to confirm the interactions among Xa1, OsERF101 and two TAL effectors. We created OsERF101-overexpressing and knockout mutant lines in rice and identified genes differentially regulated in these lines, many of which are predicted to be involved in the regulation of response to stimulus. Xa1 interacts in the nucleus with the TAL effectors and OsERF101 via the BED domain. Unexpectedly, both the overexpression and the knockout lines of OsERF101 displayed Xa1-dependent, enhanced resistance to an incompatible Xoo strain. Different sets of genes were up- or downregulated in the overexpression and knockout lines. Our results indicate that OsERF101 regulates the recognition of TAL effectors by Xa1, and functions as a positive regulator of Xa1-mediated immunity. Furthermore, an additional Xa1-mediated immune pathway is negatively regulated by OsERF101.

Incorporation of Two Bacterial Blight Resistance Genes into the Popular Rice Variety, Ranidhan through Marker-Assisted Breeding

The bacterial blight (BB) disease of rice is a major disease that reduces yield heavily in susceptible varieties. Ranidhan is a late maturing popular rice variety but shows high susceptibility to the disease. Two BB resistance genes were transferred into the variety through a marker-assisted backcross breeding approach. Tightly linked molecular markers were deployed to track the BB resistance genes in the plants carrying the target genes in each backcross generation. Foreground screening detected 17, 16 and 15 progenies to carry the 3 BB resistance genes in BC1F1, BC2F1 and BC3F1 generations, respectively. The selected BC3F1 plant was selfed and three different combinations of BB resistance genes were tracked in homozygous state in seven BC3F2 plants. The pyramided lines carrying three resistance genes in homozygous conditions were evaluated for BB disease resistance by inoculating with eight virulent Xoo strains. Five pyramided lines carrying two resistance gene combinations (Xa21+xa13 and Xa21+xa5) exhibited enhanced resistance against the BB pathogens. The disease resistance was in the order of Xa21+xa5 < Xa21+xa13 < xa13+xa5 gene combinations in conferring the resistance. The developed pyramided lines were similar to the recipient parent for the majority of the important agro-morphologic and grain quality traits.

DRR Dhan 58, a Seedling Stage Salinity Tolerant NIL of Improved Samba Mahsuri Shows Superior Performance in Multi-location Trials

BackgroundImproved Samba Mahsuri (ISM) is an elite, high-yielding, bacterial blight resistant, fine-grained rice variety with low glycaemic index. It is highly sensitive to salt stress, particularly at seedling stage, which significantly reduces its yield potential in coastal areas. A salinity tolerant QTL, Saltol, associated with seedling stage tolerance was previously mapped on chromosome 1 (10.6–11.5 Mb) from the Indian landrace, Pokkali and is effective in different genetic backgrounds. The objective of this study was to enhance salinity tolerance of ISM by incorporating the Saltol QTL through marker-assisted backcross breeding using the breeding line, FL478 (Pokkali/IR29).ResultsForeground selection was carried out at each generation using five Saltol-specific markers and three bacterial blight resistance genes, Xa21, xa13 and xa5. Background selection was conducted using 66 well distributed polymorphic SSR markers and at the BC3F2 generation, a single plant with maximum recurrent parent genome recovery (95.3%) was identified and advanced to the BC3F4 generation. Based on bacterial blight resistance, seedling stage salinity tolerance and resemblance to ISM, four advanced breeding lines were selected for testing in replicated experiments near Hyderabad, India. A promising near-isogenic line, DRR Dhan 58, was evaluated in multi-location trials-coastal salinity and it showed significant salinity tolerance, resistance to bacterial blight disease, high yield and excellent grain quality during the 2019 and 2020 trials. DRR Dhan 58 was 95.1% similar to ISM based on genotyping with the 90 K SNP chip. Whole genome resequencing analysis of Pokkali and FL478 which were salinity tolerant checks, ISM and DRR Dhan 58 showed a high degree of relatedness with respect to the candidate gene loci for Saltol and OsSKC1 (Shoot K+Concentration 1).ConclusionDRR Dhan 58, possessing Saltol and three bacterial blight resistance genes (Xa21, xa13 and xa5) in the genetic background of the Indian mega-variety of rice, Samba Mahsuri, was developed for potential cultivation in areas prone to seedling stage salinity, as well as areas with endemic bacterial blight disease. This entry had a 24% yield advantage over the recurrent parent ISM under coastal saline conditions in multi-location trials and was recently released for commercial cultivation in India.

Improvement of Bacterial Blight Resistance in Two Conventionally Cultivated Rice Varieties by Editing the Noncoding Region.

xa13 is a recessive pleiotropic gene that positively regulates rice disease resistance and negatively regulates rice fertility; thus, seriously restricting its rice breeding application. In this study, CRISPR/Cas9 gene-editing technology was used to delete the Xa13 gene promoter partial sequence, including the pathogenic bacteria-inducible expression element. Rice with the edited promoter region lost the ability for pathogen-induced gene expression without affecting background gene expression in leaves and anthers, resulting in disease resistance and normal yield. The study also screened a family of disease-resistant and normal fertile plants in which the target sequence was deleted and the exogenous transgene fragment isolated in the T1 generation (transgene-free line). Important agronomic traits of the T2 generation rice were examined. T2 generation rice with/without exogenous DNA showed no statistical differences compared to the wild type in heading stage, plant height, panicles per plant, panicle length, or seed setting rate in the field. Two important conventional rice varieties, namely Kongyu131 (KY131, Geng/japonica) and Huanghuazhan (HHZ, Xian/indica), were successfully transformed, and disease-resistant and fertile materials were obtained. Currently, these are the two important conventional rice varieties in China that can be used directly for production after improvement. Expression of the Xa13 gene in the leaves of transgenic rice (KY-PD and HHZ-PD) was not induced after pathogen infection, indicating that this method can be used universally and effectively to promote the practical application of xa13, a recessive disease-resistant pleiotropic gene, for rice bacterial blight resistance. Our study on the regulation of gene expression by editing noncoding regions of the genes provides a new idea for the development of molecular design breeding in the future.

Development of Broad Spectrum and Durable Bacterial Blight Resistant Variety through Pyramiding of Four Resistance Genes in Rice

Bacterial blight (BB) disease caused by Xanthomonas oryzae pv. oryzae is a major biotic constraint on obtaining higher grain yields in rice. Marker-assisted backcross breeding (MABB) was performed by the pyramiding of Xa4, xa5, xa13 and Xa21 resistance genes in the popular variety, Ranidhan. A foreground selection in BC1F1, BC2F1, and BC3F1 progenies detected all the target genes in 12, 7 and 16 progenies by using the closely linked markers from a population size of 446, 410, and 530, respectively. The BB-positive progenies carrying the target genes with a maximal similarity to the recipient parent was backcrossed in each backcross generation. A total of 1784 BC3F2 seeds were obtained from the best BC3F1 progeny. The screening of the BC3F2 progenies for the four target genes resulted in eight plants carrying all the four target genes. A bioassay of the pyramided lines conferred very high levels of resistance to the predominant isolates of bacterial blight disease. In addition, these pyramided lines were similar to Ranidhan in 16 morpho-quality traits, namely, plant height, filled grains/panicle, panicles/plant, grain length, grain breadth, grain weight, milling, head rice recovery, kernel length after cooking, water uptake, the volume expansion ratio, gel consistency, alkali-spreading value, and the amylose content.

Marker-assisted introgression of bacterial blight resistance gene xa13 into improved CO43

Marker-assisted introgression of bacterial blight resistance gene xa13 into improved CO43