Detection and Evaluation of Blast Resistance Genes in Backbone Indica Rice Varieties from South China.

Rice blast caused by the pathogenic fungus Magnaporthe oryzae poses a significant threat to rice cultivation. The identification of robust resistance germplasm is crucial for breeding resistant varieties. In this study, we employed functional molecular markers for 10 rice blast resistance genes, namely Pi1, Pi2, Pi5, Pi9, Pia, Pid2, Pid3, Pigm, Pikh, and Pita, to assess blast resistance across 91 indica rice backbone varieties in South China. The results showed a spectrum of resistance levels ranging from highly resistant (HR) to highly susceptible (HS), with corresponding frequencies of 0, 19, 40, 27, 5, and 0, respectively. Yearly correlations in blast resistance genes among the 91 key indica rice progenitors revealed Pid2 (60.44%), Pia (50.55%), Pita (45.05%), Pi2 (32.97%), Pikh (4.4%), Pigm (2.2%), Pi9 (2.2%), and Pi1 (1.1%). Significant variations were observed in the distribution frequencies of these 10 resistance genes among these progenitors across different provinces. Furthermore, as the number of aggregated resistance genes increased, parental resistance levels correspondingly improved, though the efficacy of different gene combinations varied significantly. This study provides the initial steps toward strategically distributing varieties of resistant indica rice genotypes across South China.

Marker-assisted breeding accelerates the development of multiple-stress-tolerant rice genotypes adapted to wider environments.

Introduction: Rice, one of the major staple food crops is frequently affected by various biotic/abiotic stresses including drought, salinity, submergence, heat, Bacterial leaf blight, Brown plant hopper, Gall midge, Stem borer, Leaf folder etc. Sustained increase of yield growth is highly necessary to meet the projected demand in rice production during the year 2050. Hence, development of high yielding and multiple stress tolerant rice varieties adapted to wider environments will serve the need. Methods: A systematic MAB approach was followed to pyramid eight major QTLs/genes controlling tolerance to major abiotic/biotic stresses viz., drought (qDTY1.1 and qDTY2.1), salinity (Saltol), submergence (Sub1), bacterial leaf blight (xa13 and Xa21), blast (Pi9) and gall midge (Gm4) in the genetic background of an elite rice culture CBMAS 14065 possessing high yield and desirable grain quality traits. Two advanced backcross derivatives of CBMAS 14065 possessing different combinations of target QTLs namely #27-1-39 (qDTY1.1+qDTY2.1+Sub1+xa13+Xa21+Gm4+Pi9) and #29-2-2 (qDTY1.1+qDTY2.1+Saltol+Xa21+Gm4+Pi9) were inter-mated. Results: Inter-mated F1 progenies harboring all the eight target QTLs/genes were identified through foreground selection. Genotyping of the inter-mated F4 population identified 14 progenies possessing all eight target QTLs/genes under homozygous conditions. All the fourteen progenies were forwarded up to F8 generation and evaluated for their yield and tolerance to dehydration, salinity, submergence, blast and bacterial leaf blight. All the 14 progenies exhibited enhanced tolerance to dehydration and salinity stresses by registering lesser reduction in their chlorophyll content, relative water content, root length, root biomass etc., against their recurrent parent Improved White Ponni/CBMAS 14065. All the 14 progenies harboring Sub1 loci from FR13A exhibited enhanced survival (90 - 95%) under 2 weeks of submergence /flooding when compared to their recurrent parent CBMAS 14065 which showed 100% susceptibility The inter-mated population showed a enhanced level of resistance to bacterial leaf blight (Score = 0 to 2) against blast (Score - 0) whereas the susceptible check CO 39 and the recurrent parent CBMAS 14065 recorded high level of susceptibility (Score = 7 to 9). Conclusion or discussion: Our study demonstrated the accelerated development of multiple stress tolerant rice genotypes through marker assisted pyramiding of target QTLs/genes using tightly linked markers. These multiple stress tolerant rice lines will serve as excellent genetic stocks for field testing/variety release and also as parental lines in future breeding programs for developing climate resilient super rice varieties.

QTL detection and candidate gene identification of qCTB1 for cold tolerance in the Yunnan plateau landrace rice.

Cold stress is one of the main abiotic stresses that affects rice growth and production worldwide. Dissection of the genetic basis is important for genetic improvement of cold tolerance in rice. In this study, a new source of cold-tolerant accession from the Yunnan plateau, Lijiangxiaoheigu, was used as the donor parent and crossed with a cold-sensitive cultivar, Deyou17, to develop recombinant inbred lines (RILs) for quantitative trait locus (QTL) analysis for cold tolerance at the early seedling and booting stages in rice. In total, three QTLs for cold tolerance at the early seedling stage on chromosomes 2 and 7, and four QTLs at the booting stage on chromosomes 1, 3, 5, and 7, were identified. Haplotype and linear regression analyses showed that QTL pyramiding based on the additive effect of these favorable loci has good potential for cold tolerance breeding. Effect assessment in the RIL and BC3F3 populations demonstrated that qCTB1 had a stable effect on cold tolerance at the booting stage in the genetic segregation populations. Under different cold stress conditions, qCTB1 was fine-mapped to a 341-kb interval between markers M3 and M4. Through the combination of parental sequence comparison, candidate gene-based association analysis, and tissue and cold-induced expression analyses, eight important candidate genes for qCTB1 were identified. This study will provide genetic resources for molecular breeding and gene cloning to improve cold tolerance in rice. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-024-01488-3.

Dissecting the molecular basis of the ultra-large grain formation in rice.

The shape of rice grains not only determines the thousand-grain weight but also correlates closely with the grain quality. Here we identified an ultra-large grain accession (ULG) with a thousand-grain weight exceeding 60 g. The integrated analysis of QTL, BSA, de novo genome assembled, transcription sequencing, and gene editing was conducted to dissect the molecular basis of the ULG formation. The ULG pyramided advantageous alleles from at least four known grain-shaping genes, OsLG3, OsMADS1, GS3, GL3.1, and one novel locus, qULG2-b, which encoded a leucine-rich repeat receptor-like kinase. The collective impacts of OsLG3, OsMADS1, GS3, and GL3.1 on grain size were confirmed in transgenic plants and near-isogenic lines. The transcriptome analysis identified 112 genes cooperatively regulated by these four genes that were prominently involved in photosynthesis and carbon metabolism. By leveraging the pleiotropy of these genes, we enhanced the grain yield, appearance, and stress tolerance of rice var. SN265. Beyond showcasing the pyramiding of multiple grain size regulation genes that can produce ULG, our study provides a theoretical framework and valuable genomic resources for improving rice variety by leveraging the pleiotropy of grain size regulated genes.

Pyramiding of triple Clubroot resistance loci conferred superior resistance without negative effects on agronomic traits in Brassica napus.

Clubroot disease caused by Plasmodiophora brassicae is becoming a serious threat to rapeseed (Brassica napus) production worldwide. Breeding resistant varieties using CR (clubroot resistance) loci is the most promising solution. Using marker-assisted selection and speed-breeding technologies, we generated Brassica napus materials in homozygous or heterozygous states using CRA3.7, CRA08.1, and CRA3.2 loci in the elite parental line of the Zhongshuang11 background. We developed three elite lines with two CR loci in different combinations and one line with three CR loci at the homozygous state. In our study, we used six different clubroot strains (Xinmin, Lincang, Yuxi, Chengdu, Chongqing, and Jixi) which are categorized into three groups based on our screening results. The newly pyramided lines with two or more CR loci displayed better disease resistance than the parental lines carrying single CR loci. There is an obvious gene dosage effect between CR loci and disease resistance levels. For example, pyramided lines with triple CR loci in the homozygous state showed superior resistance for all pathogens tested. Moreover, CR loci in the homozygous state are better on disease resistance than the heterozygous state. More importantly, no negative effect was observed on agronomic traits for the presence of multiple CR loci in the same background. Overall, these data suggest that the pyramiding of triple clubroot resistance loci conferred superior resistance with no negative effects on agronomic traits in Brassica napus.

Production of grains with ultra-low heavy metal accumulation by pyramiding novel Alleles of OsNramp5 and OsLsi2 in two-line hybrid rice.

Ensuring rice yield and grain safety quality are vital for human health. In this study, we developed two-line hybrid rice (TLHR) with ultra-low grain cadmium (Cd) and arsenic (As) accumulation by pyramiding novel alleles of OsNramp5 and OsLsi2. We first generated low Cd accumulation restorer (R) lines by editing OsNramp5, OsLCD, and OsLCT in japonica and indica. After confirming that OsNramp5 was most efficient in reducing Cd, we edited this gene in C815S, a genic male sterile line (GMSL), and screened it for alleles with low Cd accumulation. Next, we generated R and GMSL lines with low As accumulation by editing OsLsi2 in a series of YK17 and C815S lines. When cultivated in soils that were heavily polluted with Cd and As, the edited R, GMSL, and TLHR plants showed significantly reduced heavy metal accumulation, while maintaining a relatively stable yield potential. This study provides an effective scheme for the safe production of grains in As- and/or Cd-polluted paddy fields.

Modern Breeding Strategies and Tools for Durable Late Blight Resistance in Potato.

Cultivated potato (Solanum tuberosum) is a major crop worldwide. It occupies the second place after cereals (corn, rice, and wheat). This important crop is threatened by the Oomycete Phytophthora infestans, the agent of late blight disease. This pathogen was first encountered during the Irish famine during the 1840s and is a reemerging threat to potatoes. It is mainly controlled chemically by using fungicides, but due to health and environmental concerns, the best alternative is resistance. When there is no disease, no treatment is required. In this study, we present a summary of the ongoing efforts concerning resistance breeding of potato against this devastating pathogen, P. infestans. This work begins with the search for and selection of resistance genes, whether they are from within or from outside the species. The genetic methods developed to date for gene mining, such as effectoromics and GWAS, provide researchers with the ability to identify genes of interest more efficiently. Once identified, these genes are cloned using molecular markers (MAS or QRL) and can then be introduced into different cultivars using somatic hybridization or recombinant DNA technology. More innovative technologies have been developed lately, such as gene editing using the CRISPR system or gene silencing, by exploiting iRNA strategies that have emerged as promising tools for managing Phytophthora infestans, which can be employed. Also, gene pyramiding or gene stacking, which involves the accumulation of two or more R genes on the same individual plant, is an innovative method that has yielded many promising results. All these advances related to the development of molecular techniques for obtaining new potato cultivars resistant to P. infestans can contribute not only to reducing losses in agriculture but especially to ensuring food security and safety.

Comparative transcriptome analysis of canola carrying a single vs stacked resistance genes against clubroot.

Pyramiding resistance genes may expand the efficacy and scope of a canola variety against clubroot (Plasmodiophora brassicae), a serious threat to canola production in western Canada. However, the mechanism(s) of multigenic resistance, especially the potential interaction among clubroot resistance (CR) genes, are not well understood. In this study, transcriptome was compared over three canola (Brassica napus L.) inbred/hybrid lines carrying a single CR gene in chromosome A03 (CRaM, Line 16) or A08 (Crr1rutb, Line 20), and both genes (CRaM+Crr1rutb, Line 15) inoculated with a field population (L-G2) of P. brassicae pathotype X, a new variant found in western Canada recently. The line16 was susceptible, while lines 15 and 20 were partially resistant. Functional annotation identified differential expression of genes (DEGs) involved in biosynthetic processes responsive to stress and regulation of cellular process; The Venn diagram showed that the partially resistant lines 15 and 20 shared 1,896 differentially expressed genes relative to the susceptible line 16, and many of these DEGs are involved in defense responses, activation of innate immunity, hormone biosynthesis and programmed cell death. The transcription of genes involved in Pathogen-Associated Molecular Pattern (PAMP)-Triggered and Effector-Triggered Immunity (PTI and ETI) was particularly up-regulated, and the transcription level was higher in line 15 (CRaM + Crr1rutb) than in line 20 (Crr1rutb only) for most of the DEGs. These results indicated that the partial resistance to the pathotype X was likely conferred by the CR gene Crr1rutb for both lines 15 and 20 that functioned via the activation of both PTI and ETI signaling pathways. Additionally, these two CR genes might have synergistic effects against the pathotype X, based on the higher transcription levels of defense-related DEGs expressed by inoculated line 15, highlighting the benefit of gene stacking for improved canola resistance as opposed to a single CR gene alone.

Resilience of Canola to Plasmodiophora brassicae (Clubroot) Pathotype 3H under Different Resistance Genes and Initial Inoculum Levels.

In this study, we explored the resilience of a clubroot resistance (CR) stacking model against a field population of Plasmodiophora brassicae pathotype 3H. This contrasts with our earlier work, where stacking CRaM and Crr1rutb proved only moderately resistant to pathotype X. Canola varieties carrying Rcr1/Crr1rutb and Rcr1 + Crr1rutb were repeatedly exposed to 3H at low (1 × 104/g soil) and high (1 × 107/g soil) initial resting spore concentrations over five planting cycles under controlled environments to mimic intensive canola production. Initially, all resistant varieties showed strong resistance. However, there was a gradual decline in resistance over time for varieties carrying only a single CR gene, particularly with Crr1rutb alone and at the high inoculum level, where the disease severity index (DSI) increased from 9% to 39% over five planting cycles. This suggests the presence of virulent pathotypes at initially low levels in the 3H inoculum. In contrast, the variety with stacked CR genes remained resilient, with DSI staying below 3% throughout, even at the high inoculum level. Furthermore, the use of resistant varieties, carrying either a single or stacked CR genes, reduced the total resting spore numbers in soil over time, while the inoculum level either increased or remained high in soils where susceptible Westar was continuously grown. Our study demonstrates greater resistance resilience for stacking Rcr1 and Crr1rutb against the field population of 3H. Additionally, the results suggest that resistance may persist even longer in fields with lower levels of inoculum, highlighting the value of extended crop rotation (reducing inoculum) alongside strategic CR-gene deployment to maximize resistance resilience.

Multiplex PCR for Early Generation Identification of Tomato Segregants Carrying Ty-2, Ty-3 and Ph-3 Resistance Alleles Against Leaf Curl and Late Blight Diseases.

Deployment of different natural disease resistance alleles is the most sustainable and eco-friendly way for multiple disease management in tomato. Diagnostic molecular markers are indispensible in this effort as they offer early generation identification of resistance alleles in an environment-independent manner. Moreover, optimized multiplex polymerase chain reaction (PCR) for detecting different disease resistance alleles in a single reaction can speed-up the selection process with cost and labour-effectiveness. Here we report the optimized multiplex detection and stacking of leaf curl disease resistance alleles Ty-2 and Ty-3 along with late blight disease resistance allele Ph-3 in tomato genotypes and F2 segregants. The triplex assay could be replaced by a duplex assay (for Ty-2 and Ty-3 resistance alleles) followed by analysis at Ph-3 locus to achieve further cost-effectiveness. We identified two plants in F2 populations derived from the Arka Samrat (F1) x Kashi Chayan combination to carry the Ty-2, Ty-3 and Ph-3 resistance alleles in homozygous condition. Early generation genotyping also allowed us to identify a few morphologically better segregants, where further marker assisted selection (MAS) should identify superior multiple disease resistant lines. Thus we advocate the utility of multiplex PCR in MAS to address multiple disease resistance breeding in tomato.

Pyramiding aphid resistance genes into the elite cowpea variety, Zaayura, using marker-assisted backcrossing.

The cowpea aphid (Aphis cracivora) is a cosmopolitan insect pest that causes economic damage on cowpea. Although the pest persists at all the growth stages of the crop, in West Africa, aphids are the only major insect pests that farmers regularly control at the vegetative stage. Thus, deploying aphid-resistant crop varieties can reduce farmers' expenditure on insecticide. The availability of different biotypes of the pest and reports of resistance breakdown necessitates pyramiding of sources of aphid resistance to develop a more robust genotype for durable resistance. Two aphid-resistance genes, sourced from SARC-1-57-2 and IT97K-556-6, were introgressed through gene pyramiding technique into a farmers' preferred cowpea variety, Zaayura, using marker-assisted backcrossing. A simple sequence repeat (SSR) marker, CP 171F/172R, and an allele-specific single nucleotide polymorphism (SNP) marker, 1_0912, were used for foreground selection of the SARC-1-57-2 and IT97K-556-6 aphid resistance genes, respectively. A stepwise backcross approach was used to introgress the major aphid resistance QTL (QAc-vu7.1) from IT97K-556-6 into Zaayura using the marker 1_0912 coupled with intermittent screening under artificial aphid infestation. After the fourth backcross generation, three heterozygous BC4F1 of Zaayura/TT97K-556-6 were intercrossed to Zaayura Pali to develop intercross F1 (ICF1). Three true ICF1 hybrids allowed to self to produce ICF2. Five (5) out of 48 ICF2 plants which were genotyped with the two foreground markers had the two aphid resistance genes fixed in the double homozygous dominant state. For background selection, out of 192 allele-specific markers screened, only 47 polymorphic markers were identified and used for the background analysis of the pyramided lines. The recurrent parent genome recovery ranged from 72 to 93.8 %. ICF2_Zaa/556/SARC-P6 had the highest recurrent parent genome and the least heterozygosity among the five improved lines. The five pyramided lines showed superior resistance under artificial aphid infestation as compared to the two donor parents with damage scores ranging from 2.0 to 2.3. On the field, however, there were no significant differences between the pyramided lines and their recurrent parent for all the agronomic traits measured except for grain yield. The pyramided lines do not only stand the chance of being released as new varieties but are also valuable genetic resources for other breeding programs that seek to improve cowpea for aphid resistance.

Deleterious Effects of Heat Stress on the Tomato, Its Innate Responses, and Potential Preventive Strategies in the Realm of Emerging Technologies.

The tomato is a fruit vegetable rich in nutritional and medicinal value grown in greenhouses and fields worldwide. It is severely sensitive to heat stress, which frequently occurs with rising global warming. Predictions indicate a 0.2 °C increase in average surface temperatures per decade for the next three decades, which underlines the threat of austere heat stress in the future. Previous studies have reported that heat stress adversely affects tomato growth, limits nutrient availability, hammers photosynthesis, disrupts reproduction, denatures proteins, upsets signaling pathways, and damages cell membranes. The overproduction of reactive oxygen species in response to heat stress is toxic to tomato plants. The negative consequences of heat stress on the tomato have been the focus of much investigation, resulting in the emergence of several therapeutic interventions. However, a considerable distance remains to be covered to develop tomato varieties that are tolerant to current heat stress and durable in the perspective of increasing global warming. This current review provides a critical analysis of the heat stress consequences on the tomato in the context of global warming, its innate response to heat stress, and the elucidation of domains characterized by a scarcity of knowledge, along with potential avenues for enhancing sustainable tolerance against heat stress through the involvement of diverse advanced technologies. The particular mechanism underlying thermotolerance remains indeterminate and requires further elucidatory investigation. The precise roles and interplay of signaling pathways in response to heat stress remain unresolved. The etiology of tomato plants' physiological and molecular responses against heat stress remains unexplained. Utilizing modern functional genomics techniques, including transcriptomics, proteomics, and metabolomics, can assist in identifying potential candidate proteins, metabolites, genes, gene networks, and signaling pathways contributing to tomato stress tolerance. Improving tomato tolerance against heat stress urges a comprehensive and combined strategy including modern techniques, the latest apparatuses, speedy breeding, physiology, and molecular markers to regulate their physiological, molecular, and biochemical reactions.

[Genetic dissection of rice resistance to bacterial blight].

Bacterial blight, a major disease in rice, poses a serious impact on rice production. In this study, a doubled haploid (DH) population derived from a cross between the introduced japonica cultivar 'Maybelle' and the indica landrace 'Baiyeqiu' was used to investigate the pathogenicity of four pathogen races causing bacterial blight. The results showed that the pathogenicity of all the pathogen races exhibited continuous, transgressive distribution in the DH population. Moreover, strong correlations existed between every two pathogen races, with the correlation coefficients ranging from 0.3 to 0.6. A total of 12 quantitative trait loci (QTLs) distributed on chromosomes 1, 2, 3, 5, 6, 7, 9, and 12 were detected for rice bacterial blight, explaining 4.95% to 16.05% of the phenotype. Among these QTLs, a major QTL located in the interval RM6024-RM163 on chromosome 5 was detected in three pathogen races. In addition, the pyramiding of the positive alleles can apparently improve the rice resistance to bacterial blight. This study is of great significance for broadening the genetic resources with resistance to bacterial blight in China.

Gene pyramiding of ZmGLK36 and ZmGDIα-hel for rough dwarf disease resistance in maize.

Maize rough dwarf disease (MRDD) caused by pathogenic viruses in the genus Fijivirus in the family Reoviridae is one of the most destructive diseases in maize. The pyramiding of effective resistance genes into maize varieties is a potential approach to reduce the damage resulting from the disease. Two major quantitative trait loci (QTLs) (qMrdd2 and qMrdd8) have been previously identified. The resistance genes ZmGLK36 and ZmGDIα-hel have also been cloned with the functional markers Indel-26 and IDP25K, respectively. In this study, ZmGLK36 and ZmGDIα-hel were introgressed to improve MRDD resistance of maize lines (Zheng58, Chang7-2, B73, Mo17, and their derived hybrids Zhengdan958 and B73 × Mo17) via marker-assisted selection (MAS). The converted lines and their derived hybrids, carrying one or two genes, were evaluated for MRDD resistance using artificial inoculation methods. The double-gene pyramiding lines and their derived hybrids exhibited increased resistance to MRDD compared to the monogenic lines and the respective hybrids. The genetic backgrounds of the converted lines were highly similar (90.85-98.58%) to the recurrent parents. In addition, agronomic trait evaluation demonstrated that pyramiding lines with one or two genes and their derived hybrids were not significantly different from the recurrent parents and their hybrids under nonpathogenic stress, including period traits (tasseling, pollen shedding, and silking), yield traits (ear length, grain weight per ear and 100-kernel weight) and quality traits (protein and starch content). There were differences in plant architecture traits between the improved lines and their hybrids. This study illustrated the successful development of gene pyramiding for improving MRDD resistance by advancing the breeding process. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-024-01466-9.

The Function of SD1 on Shoot Length and its Pyramiding Effect on Shoot Length and Plant Height in Rice (Oryza sativa L.).

Strong seedling vigor is imperative to achieve stable seedling establishment and enhance the competitiveness against weeds in rice direct seeding. Shoot length (SL) is one of the important traits associated with seedling vigor in rice, but few genes for SL have been cloned so far. In the previous study, we identified two tightly linked and stably expressed QTLs for SL, qSL-1f and qSL-1d by genome-wide association study, and cloned the causal gene (LOC_Os01g68500) underlying qSL-1f. In the present study, we identify LOC_Os01g66100 (i.e. the semidwarf gene SD1), a well-known gene controlling plant height (PH) at the adult-plant stage, as the causal gene underlying qSL-1d through gene-based haplotype analysis and knockout transgenic verification. By measuring the phenotypes (SL and PH) of various haplotypes of the two genes and their knockout lines, we found SD1 and LOC_ Os01g68500 controlled both SL and PH, and worked in the same direction, which provided the directly genetic evidence for a positive correlation between SL and PH combined with the analysis of SL and PH in the diverse natural population. Moreover, the knockout transgenic experiments suggested that SD1 had a greater effect on PH compared with LOC_ Os01g68500, but no significant difference in the effect on SL. Further investigation of the pyramiding effects of SD1 and LOC_Os01g68500 based on their haplotype combinations suggested that SD1 may play a dominant role in controlling SL and PH when the two genes coexist. In this study, the effect of SD1 on SL at the seedling stage is validated. In total, two causal genes, SD1 and LOC_ Os01g68500, for SL are cloned in our studies, which controlled both SL and PH, and the suitable haplotypes of SD1 and LOC_ Os01g68500 are beneficial to achieve the desired SL and PH in different rice breeding objectives. These results provide a new clue to develop rice varieties for direct seeding and provide new genetic resources for molecular breeding of rice with suitable PH and strong seedling vigor.

Pyramiding D-lactate dehydrogenase with the glyoxalase pathway enhances abiotic stress tolerance in plants.

Methylglyoxal is a common cytotoxic metabolite produced in plants during multiple biotic and abiotic stress. To mitigate the toxicity of MG, plants utilize the glyoxalase pathway comprising glyoxalase I (GLYI), glyoxalase II (GLYII), or glyoxalase III (GLYIII). GLYI and GLYII are the key enzymes of glyoxalase pathways that play an important role in abiotic stress tolerance. Earlier research showed that MG level is lower when both GLYI and GLYII are overexpressed together, compared to GLYI or GLYII single gene overexpressed transgenic plants. D-lactate dehydrogenase (D-LDH) is an integral part of MG detoxification which metabolizes the end product (D-lactate) of the glyoxalase pathway. In this study, two Arabidopsis transgenic lines were constructed using gene pyramiding technique: GLYI and GLYII overexpressed (G-I + II), and GLYI, GLYII, and D-LDH overexpressed (G-I + II + D) plants. G-I + II + D exhibits lower MG and D-lactate levels and enhanced abiotic stress tolerance than the G-I + II and wild-type plants. Further study explores the stress tolerance mechanism of G-I + II + D plants through the interplay of different regulators and plant hormones. This, in turn, modulates the expression of ABA-dependent stress-responsive genes like RAB18, RD22, and RD29B to generate adaptive responses during stress. Therefore, there might be a potential correlation between ABA and MG detoxification pathways. Furthermore, higher STY46, GPX3, and CAMTA1 transcripts were observed in G-I + II + D plants during abiotic stress. Thus, our findings suggest that G-I + II + D has significantly improved MG detoxification, reduced oxidative stress-induced damage, and provided a better protective mechanism against abiotic stresses than G-I + II or wild-type plants.

Pyramiding of Low Chalkiness QTLs Is an Effective Way to Reduce Rice Chalkiness.

Rice chalkiness is a key limiting factor of high-quality rice. The breeding of low chalkiness varieties has always been a challenging task due to the complexity of chalkiness and its susceptibility to environmental factors. In previous studies, we identified six QTLs for the percentage of grain chalkiness (PGC), named qPGC5, qPGC6, qPGC8.1, qPGC8.2, qPGC9 and qPGC11, using single-segment substitution lines (SSSLs) with genetic background of Huajingxian 74 (HJX74). In this study, we utilized the six low chalkiness QTLs to develop 17 pyramiding lines with 2-4 QTLs. The results showed that the PGC decreased with the increase of QTLs in the pyramiding lines. The pyramiding lines with 4 QTLs significantly reduced the chalkiness of rice and reached the best quality level. Among the six QTLs, qPGC5 and qPGC6 showed greater additive effects and were classified as Group A, while the other four QTLs showed smaller additive effects and were classified as Group B. In pyramiding lines, although the presence of epistasis, additivity remained the main component of QTL effects. qPGC5 and qPGC6 showed stronger ability to reduce rice chalkiness, particularly in the environment of high temperature (HT) in the first cropping season (FCS). Our research demonstrates that by pyramiding low chalkiness QTLs, it is feasible to develop the high-quality rice varieties with low chalkiness at the best quality level even in the HT environment of FCS.

Gene pyramiding for boosted plant growth and broad abiotic stress tolerance.

Abiotic stresses such as salinity, heat and drought seriously impair plant growth and development, causing a significant loss in crop yield and ornamental value. Biotechnology approaches manipulating specific genes prove to be effective strategies in crop trait modification. The Arabidopsis vacuolar pyrophosphatase gene AVP1, the rice SUMO E3 ligase gene OsSIZ1 and the cyanobacterium flavodoxin gene Fld have previously been implicated in regulating plant stress responses and conferring enhanced tolerance to different abiotic stresses when individually overexpressed in various plant species. We have explored the feasibility of combining multiple favourable traits brought by individual genes to acquire superior plant performance. To this end, we have simultaneously introduced AVP1, OsSIZ1 and Fld in creeping bentgrass. Transgenic (TG) plants overexpressing these three genes performed significantly better than wild type controls and the TGs expressing individual genes under both normal and various abiotic stress conditions, exhibited significantly enhanced plant growth and tolerance to drought, salinity and heat stresses as well as nitrogen and phosphate starvation, which were associated with altered physiological and biochemical characteristics and delicately fine-tuned expression of genes involved in plant stress responses. Our results suggest that AVP1, OsSIZ1 and Fld function synergistically to regulate plant development and plant stress response, leading to superior overall performance under both normal and adverse environments. The information obtained provides new insights into gene stacking as an effective approach for plant genetic engineering. A similar strategy can be extended for the use of other beneficial genes in various crop species for trait modifications, enhancing agricultural production.

Pyramiding BPH genes in rice maintains resistance against the brown planthopper under climate change.

BACKGROUND: Nilaparvata lugens (brown planthopper; BPH) is a significant rice pest in Asia, causing substantial yield losses. Pyramiding BPH resistance genes with diverse resistance traits into rice cultivars is an effective strategy for pest management. However, the response of pyramiding combinations to environmental changes remains unclear. To address this knowledge gap, we investigated three pyramiding rice lines (BPH2 + 32, BPH9 + 32, and BPH18 + 32) in the context of varying climate change conditions, ensuring sufficient N. lugens-rice interactions. Thus, we set three environmental conditions [30/25 °C (day/night) with 500 ppm CO2 concentration, 32/27 °C (day/night) with 600 ppm CO2 concentration, and 35/30 °C (day/night) with 1000 ppm CO2 concentration]. RESULTS: All three pyramiding rice lines maintained the insect resistant ability under the three environmental settings. In particular, the BPH18 + 32 rice line exhibited stronger antibiotic and antixenosis effects against N. lugens. In addition, BPH18 + 32 rice line had better shoot resilience under N. lugens infestation, whereas the performance of the other two selected pyramiding rice lines varied. Thus, although BPH2, BPH9, and BPH18 represent three alleles at the same locus, their resistance levels against N. lugens may vary under distinct climate change scenarios, as evidenced by the performance of N. lugens on the three pyramiding rice lines. CONCLUSION: Our findings indicate that all three tested pyramiding rice lines maintained their insect resistance in the face of diverse climate change scenarios. However, these lines exhibited varied repellent responses and resilience capacities in response to climate change. Thus, the combination of pyramiding genes needs to be considered for future breeding programs. © 2023 Society of Chemical Industry.

Dissecting the major genetic components underlying cotton lint development.

Numerous genetic loci and several functionally characterized genes have been linked to determination of lint percentage (lint%), one of the most important cotton yield components, but we still know little about the major genetic components underlying lint%. Here, we first linked the genetic loci containing MYB25-like_At and HD1_At to the fiberless seed trait of 'SL1-7-1' and found that MYB25-like_At and HD1_At were very lowly expressed in 'SL1-7-1' ovules during fiber initiation. We then dissected the genetic components involved in determination of lint% using segregating populations derived from crosses of fuzzless mutants and intermediate segregants with different lint%, which not only confirmed the HD1_At locus but identified the HD1_Dt locus as being the major genetic components contributing to fiber initiation and lint%. The segregating populations also allowed us to evaluate the relative contributions of MYB25-like_At, MYB25-like_Dt, HD1_At, and HD1_Dt to lint%. Haplotype analysis of an Upland cotton (Gossypium hirsutum) population with 723 accessions (including 81 fuzzless seed accessions) showed that lint% of the accessions with the LP allele (higher lint%) at MYB25-like_At, MYB25-like_Dt, or HD1_At was significantly higher than that with the lp allele (lower lint%). The lint% of the Upland cotton accessions with 3 or 4 LP alleles at MYB25-like and HD1 was significantly higher than that with 2 LP alleles. The results prompted us to propose a strategy for breeding high-yielding cotton varieties, i.e. pyramiding the LP alleles of MYB25-like and HD1 with new lint% LP alleles without negative impact on seed size and fiber quality.