Progress on Salt Tolerance in Brassica napus.

In China, saline-alkali lands constitute 5.01% of the total land area, having a significant impact on both domestic and international food production. Rapeseed (Brassica napus L.), as one of the most important oilseed crops in China, has garnered considerable attention due to its potential adaptability to saline conditions. Breeding and improving salt-tolerant varieties is a key strategy for the effective utilization of saline lands. Hence, it is important to conduct comprehensive research into the adaptability and salt tolerance mechanisms of Brassica napus in saline environments as well as to breed novel salt-tolerant varieties. This review summarizes the molecular mechanism of salt tolerance, physiological and phenotypic indexes, research strategies for the screening of salt-tolerant germplasm resources, and genetic engineering tools for salt stress in Brassica napus. It also introduces various agronomic strategies for applying exogenous substances to alleviate salt stress and provide technological tools and research directions for future research on salt tolerance in Brassica napus.

Fine mapping of the panicle length QTL qPL5 in rice.

Panicle length is a crucial trait tightly associated with spikelets per panicle and grain yield in rice. To dissect the genetic basis of panicle length, a population of 161 recombinant inbred lines (RILs) was developed from the cross between an aus variety Chuan 7 (C7) and a tropical Geng variety Haoboka (HBK). C7 has a panicle length of 30 cm, 7 cm longer than that of HBK, and the panicle length was normally distributed in the RIL population. A total of six quantitative trait loci (QTLs) for panicle length were identified, and single QTLs explained the phenotypic variance from 4.9 to 18.1%. Among them, three QTLs were mapped to the regions harbored sd1, DLT, and Ehd1, respectively. To validate the genetic effect of a minor QTL qPL5, a near-isogenic F2 (NIF2) population segregated at qPL5 was developed. Interestingly, panicle length displayed bimodal distribution, and heading date also exhibited significant variation in the NIF2 population. qPL5 accounted for 66.5% of the panicle length variance. The C7 allele at qPL5 increased panicle length by 2.4 cm and promoted heading date by 5 days. Finally, qPL5 was narrowed down to an 80-kb region flanked by markers M2197 and M2205 using a large NIF2 population of 7600 plants. LOC_Os05g37540, encoding a phytochrome signal protein whose homolog in Arabidopsis enlarges panicle length, is regarded as the candidate gene because a single-nucleotide mutation (C1099T) caused a premature stop codon in HBK. The characterization of qPL5 with enlarging panicle length but promoting heading date makes its great value in breeding early mature varieties without yield penalty in rice. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-024-01443-2.

A Pseudo-near isogenic F2 population strategy for rapid QTL cloning.

Near isogenic F2 (NIF2) population frequently developed by conventional backcross has dramatically contributed to QTL identification in plants. Developing such a NIF2 population is time-consuming. Thus, it is urgent to rapidly produce a NIF2 population for QTL cloning. Here, we proposed a rapid QTL cloning strategy by generating a Pseudo-near isogenic F2 population (Pseudo-NIF2), which segregates at the target QTL but is fixed at other QTLs for the target trait. Nineteen QTLs for GL, GW, and TGW were detected in the F2 population from the cross between Zhenshan 97 and Egy316. To verify the efficiency of Pseudo-NIF2 in QTL quick cloning, the novel moderate QTL qGL10.1 which explained 9.1% and 5.6% of grain length variation in F2 and F2:3 populations was taken as an example. An F2 plant (F2-120), which segregated at qGL10.1 but fixed at other 8 QTLs for grain length, was screened to generate a Pseudo-NIF2 population by selfing cross. In the Pseudo-NIF2 population, the segregation ratio of plants with long grains to short grains fits 3:1, indicating that one gene controlled the variation of grain length. Based on the Pseudo-NIF2 and its progeny, qGL10.1 was fine mapped to a 19.3-kb region, where a gene OsMADS56 was verified as the candidate by functional polymorphism between parental alleles. Pseudo-NIF2 strategy is a rapid way for QTL cloning, which saves 3 to 4 cropping seasons compared to the conventional way. Applying the method for cloning QTL with moderate or major effects is promising. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-023-01408-x.

Suppressing a phosphohydrolase of cytokinin nucleotide enhances grain yield in rice.

One-step and two-step pathways are proposed to synthesize cytokinin in plants. The one-step pathway is mediated by LONELY GUY (LOG) proteins. However, the enzyme for the two-step pathway remains to be identified. Here, we show that quantitative trait locus GY3 may boost grain yield by more than 20% through manipulating a two-step pathway. Locus GY3 encodes a LOG protein that acts as a 5'-ribonucleotide phosphohydrolase by excessively consuming the cytokinin precursors, which contrasts with the activity of canonical LOG members as phosphoribohydrolases in a one-step pathway. The residue S41 of GY3 is crucial for the dephosphorylation of iPRMP to produce iPR. A solo-LTR insertion within the promoter of GY3 suppressed its expression and resulted in a higher content of active cytokinins in young panicles. Introgression of GY302428 increased grain yield per plot by 7.4% to 16.3% in all investigated indica backgrounds, which demonstrates the great value of GY302428 in indica rice production.

Assembly of yield heterosis of an elite rice hybrid is promising by manipulating dominant quantitative trait loci.

In the past, rice hybrids with strong heterosis have been obtained empirically, by developing and testing thousands of combinations. Here, we aimed to determine whether heterosis of an elite hybrid could be achieved by manipulating major quantitative trait loci. We used 202 chromosome segment substitution lines from the elite hybrid Shanyou 63 to evaluate single segment heterosis (SSH) of yield per plant and identify heterotic loci. All nine detected heterotic loci acted in a dominant fashion, and no SSH exhibited overdominance. Functional alleles of key yield-related genes Ghd7, Ghd7.1, Hd1, and GS3 were dispersed in both parents. No functional alleles of three investigated genes were expressed at higher levels in the hybrids than in the more desirable parents. A hybrid pyramiding eight heterotic loci in the female parent Zhenshan 97 background had a comparable yield to Shanyou 63 and much higher yield than Zhenshan 97. Five hybrids pyramiding eight or nine heterotic loci in the combined parental genome background showed similar yield performance to that of Shanyou 63. These results suggest that dominance underlying functional complementation is an important contributor to yield heterosis and that heterosis assembly might be successfully promised by manipulating several major dominant heterotic loci.

Combinations of Ghd7, Ghd8, and Hd1 determine strong heterosis of commercial rice hybrids in diverse ecological regions.

Heterosis of grain yield is closely associated with heading date in crops. Gene combinations of the major heading date genes Ghd7, Ghd8, and Hd1 play important roles in enhancing grain yield and adaptation to ecological regions in rice. However, the predominant three-gene combinations for a specific ecological region remain unclear in both three-line and two-line hybrids. In this study, we sequenced these three genes of 50 cytoplasmic male sterile/maintainer lines, 31 photo-thermo-sensitive genic male sterile lines, and 109 restorer lines. Sequence analysis showed that hybrids carrying strong functional alleles of Ghd7 and Hd1 and non-functional Ghd8 are predominant in three-line hybrids and are recommended for rice production in the subtropics around 30°N/S. Hybrids carrying strong functional Ghd7 and Ghd8 and non-functional Hd1 are predominant in two-line hybrids and are recommended for low latitude areas around 23.5°N/S rich in photothermal resources. Hybrids carrying strong functional Ghd7 and Ghd8 and functional Hd1 were not identified in commercial hybrids in the middle and lower reaches of the Yangtze River, but they have high yield potential in tropical regions because they have the strongest photoperiod sensitivity. Based on these findings, two genic sterile lines, Xiangling 628S and C815S, whose hybrids often head very late, were diagnosed with these three genes, and Hd1 was targeted to be knocked out in Xiangling 628S and replaced with hd1 in C815S. The hybrids developed from both modified sterile lines in turn had appropriate heading dates and significantly improved grain yield. This study provides new insights for breeding design to develop hybrids for various regions.

The transcriptional repressor OsPRR73 links circadian clock and photoperiod pathway to control heading date in rice.

The phase transition from vegetative to reproductive growth is triggered by internal and external signals that participate in circadian clock in plants. We identified a rice floral inhibitor OsPRR73 encoding a CONSTANS protein. Overexpression of OsPRR73 resulted in late heading under both long-day (LD) and short-day (SD) conditions. Knockout mutants led to early heading under LD conditions but no change under SD. OsPRR73 mRNA accumulated at noon and exhibited a robust oscillation under constant light (LL) and constant darkness (DD) conditions. OsPRR73 overexpression exerted negative feedback on endogenous OsPRR73 expression and altered diurnal expressions of key flowering genes and circadian clock genes. OsPRR73 bound to the promoters of the floral gene Ehd1 and the circadian gene OsLHY, and significantly suppressed their expression at dawn. In LL and DD, the oscillatory patterns of the circadian genes OsLHY, OsTOC1, OsGI and OsELF3 were varied in OsPRR73OX and osprr73 mutants. OsPRR73 expression was decreased in osphyb mutants, and overexpression of OsPRR73 complemented the early heading date phenotype of osphyb, indicating OsPRR73 works downstream of OsPhyB. Therefore, OsPRR73 is involved in a feedback loop of the rice clock and connects the photoperiod flowering pathway by binding to the Ehd1 promoter in rice.

Global analysis of CCT family knockout mutants identifies four genes involved in regulating heading date in rice.

Many genes encoding CCT domain-containing proteins regulate flowering time. In rice (Oryza sativa), 41 such genes have been identified, but only a few have been shown to regulate heading date. Here, to test whether and how additional CCT family genes regulate heading date in rice, we classified these genes into five groups based on their diurnal expression patterns. The expression patterns of genes in the same subfamily or in close phylogenetic clades tended to be similar. We generated knockout mutants of the entire gene family via CRISPR/Cas9. The heading dates of knockout mutants of only 4 of 14 genes previously shown to regulate heading date were altered, pointing to functional redundancy of CCT family genes in regulating this trait. Analysis of mutants of four other genes showed that OsCCT22, OsCCT38, and OsCCT41 suppress heading under long-day conditions and promote heading under short-day conditions. OsCCT03 promotes heading under both conditions and upregulates the expression of Hd1 and Ehd1, a phenomenon not previously reported for other such genes. To date, at least 18 CCT domain-containing genes involved in regulating heading have been identified, providing diverse, flexible gene combinations for generating rice varieties with a given heading date.

Rapid identification of candidate genes controlling male-sterility in Foxtail millet (Setaria italica).

Photo-(thermo-) sensitive genic male-sterile line is the key component of two-line hybridization system in foxtail millet (Setaria italica), but the genetic basis of male sterility in most male-sterile lines is still unclear. In the present study, a large F2 population was developed derived from a cross between the photo-(thermo-) sensitive male-sterile line A2 and the fertile-line 1484-5. Thirty plants with extreme high and extreme low fertility were selected from the population to construct a sterile DNA pool and a fertile DNA pool, respectively. Sequencing both DNA pools and data analysis revealed that two QTLs conferred male-sterility, qSiMS6.1 with a major effect and qSiMS6.2 with a minor effect, on chromosome 6. Both QTLs exhibited complete dominance. The major QTL, qSiMS6.1, was delimited to a 186-kb interval between the markers SiM20 and SiM9 by the joint analysis of QTL-seq and QTL mapping with SSR and structure variation markers. Millet_GLEAN_10020454 in this region is the most likely candidate gene for qSiMS6.1 since it is predicted to encode a male-sterile 5 like protein. These results lay a solid foundation for qSiMS6.1 cloning and provided gene resources for breeding new male-sterile lines. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-021-01269-2.

Impact of diverse background interferences on the alternating trilinear decomposition modeling of excitation-emission matrix fluorescence data acquired from different sample sources.

Alternating trilinear decomposition (ATLD) method enables the qualitative and quantitative analysis of excitation-emission matrix fluorescence (EEMF) data acquired from complex samples. However, the impact of diverse background interferences from different sample sources on the performances of ATLD method has never been lucubrated. In this work, simulated and real EEMF data sets from different sample sources with diverse background interferences were collected and subjected to ATLD analysis. The performances of ATLD modeling individual and global EEMF data sets were comprehensively compared in terms of the resolved spectral profiles and quantitative results. It was found that ATLD method can use the same set of calibration samples to resolve and quantify multiple components of interest in multiple complex systems with diverse background interferences, regardless of individual or global modeling. The results revealed that the qualitative and quantitative results provided by ATLD method were affected neither by diversity of background interferences nor by data merging as long as the acquired EEMF data sets conform to the trilinear component model. This property of ATLD method can enrich the "second-order advantage", i.e. the term "unknown interferences" in the concept of "second-order advantage" refers to not only constant background interferences but also diverse background interferences, which will be certain to further expand the practicality of ATLD method in complex sample analysis, especially in the field of fluorescence spectroscopy.

CCT domain-containing genes in cereal crops: flowering time and beyond.

KEY MESSAGE: The review summarizes the functions of the plant special transcription factors CCT family genes in multiple traits and discusses the molecular breeding strategies with CCT family genes in the future. Plants integrate circadian clock and external signals such as temperature and photoperiod to synchronize flowering with seasonal environmental changes. This process makes cereal crops including short-day crops, such as rice and maize, and long-day crops, such as wheat and barley, better adapt to varied growth zones from temperate to tropical regions. CCT family genes involve circadian clock and photoperiodic flowering pathways and help plants set a suitable flowering time to produce offspring. Beyond the flowering time, CCT family genes in cereal crops are associated with biomass and grain yield. Moreover, recent studies showed that they also associate with photosynthesis, nutrition use efficiency and stress tolerance. Here, we systematically review the progress in functional characterization of CCT family genes in flowering, geographical adaptation and grain yield formation, raise the core questions related to their molecular mechanisms and discuss how to practice them in genetic improvement in cereal crops by combining gene diagnosis and top-level design.

Gene diagnosis and targeted breeding for blast-resistant Kongyu 131 without changing regional adaptability.

The fungus Magnaporthe oryzae threatens the rice production of Kongyu 131 (KY131), a leading japonica variety in Northeast China. In this study, two rice lines, KP1 and KP2-Hd1, were obtained by introgressing the blast resistance genes Pi1 and Pi2 into KY131, respectively. However, both lines headed later than KY131. RICE60K SNP array analysis showed that Hd1 closely linked to Pi2 was introgressed into KP2-Hd1, and the linkage drag of Hd1 was broken by recombination. On the other hand, no known flowering genes were introgressed into KP1. Gene diagnosis by resequencing six flowering genes showed that KP1 carried functional Hd16 and Ghd8 alleles. Due to its suppression role in heading under long-day conditions, Ghd8 was chosen as the target for gene editing to disrupt its function. Four sgRNAs targeting different sites within Ghd8 were utilized to induce large-deletion mutations, which were easy to detect via agarose gel electrophoresis. All the ghd8-mutated KP1 lines were resistant to rice blast disease and headed earlier than the control KP1, even than KY131, under natural long-day conditions, which ensures its growth in Northeast China. This study confirmed that a combination of gene diagnosis and targeted gene editing is a highly efficient way to quickly eliminate undesired traits in a breeding line.

A Rice Genetic Improvement Boom by Next Generation Sequencing.

Rice (Oryza sativa L.) is a staple food crop for people worldwide, and a key goal has been to increase its grain yield. An increasing population that relies on a decreasing level of farmland has rendered the traditional method for the isolation and use of genetic loci in rice breeding unsatisfactory. Recently, the rapid development in next generation sequencing (NGS) has boosted the number of genome sequences for hundreds to thousands of rice varieties. A MutMap strategy and bulk segregation analysis (BSA) has been developed to directly identify candidate genes based on NGS. The genome-wide association analysis (GWAS) has become a commonly used approach toward identifying the genetic loci and candidate genes for several traits that are closely associated with grain yield. The Multi-parent Advanced Generation Inter-Cross population (MAGIC) is introduced here to discuss potential applications for mapping QTLs for rice varietal development. These strategies broaden the capacity of functional gene identification and its application as a complementary method to insert mutants that comprise T-DNA and transposons. High-throughput SNP analysis platforms, such as the SNP array, provide novel strategies for genomic-assisted selections (GAS) for rice genetic improvements. Moreover, accurate genome sequence information enables genome editing for the utilization of key recessive genes that control important agronomic traits. This review summarizes how NGS accelerates rice genetic improvements through the identification and utilization of key functional genes that regulate agronomic traits.

Alternative functions of Hd1 in repressing or promoting heading are determined by Ghd7 status under long-day conditions.

Previous studies suggested that Hd1 promoted heading under short-day conditions (SD) and delayed heading under long-day conditions (LD). However in this study, Hd1 was demonstrated to consistently promote heading date in Zhenshan 97 (ZS97) background by upregulating Ehd1, Hd3a and RFT1 expression under both SD and LD. While the high photoperiod sensitivity of Hd1 was observed in Minghui 63 (MH63) background, with heading being suppressed in LD but promoted in SD. Comparative analysis of two sets of near isogenic lines of Hd1 in MH63 and ZS97 backgrounds indicated that the alternative functions of Hd1 in promoting or suppressing heading under LD are dependent on the previously cloned flowering repressor gene Ghd7. The interaction between proteins Ghd7 and Hd1 occurred through binding of the CCT domain of Ghd7 to the transcription-activating domain of Hd1, resulting in suppression of Ehd1 and florigen gene expression. The involvement of the transcription-activating domain of Hd1 in this protein-protein interaction probably blocked or weakened its transcriptional activity. These findings suggest that Hd1 alone essentially acts as a promoter of heading date, and the protein interaction between Ghd7 and Hd1 determines photoperiod sensitivity and integrated Hd1-mediated and Ehd1-mediated flowering pathways in rice.

[The application of genome editing in identification of plant gene function and crop breeding].

Plant genome can be modified via current biotechnology with high specificity and excellent efficiency. Zinc finger nucleases (ZFN), transcription activator-like effector nucleases (TALEN) and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) system are the key engineered nucleases used in the genome editing. Genome editing techniques enable gene targeted mutagenesis, gene knock-out, gene insertion or replacement at the target sites during the endogenous DNA repair process, including non-homologous end joining (NHEJ) and homologous recombination (HR), triggered by the induction of DNA double-strand break (DSB). Genome editing has been successfully applied in the genome modification of diverse plant species, such as Arabidopsis thaliana, Oryza sativa, and Nicotiana tabacum. In this review, we summarize the application of genome editing in identification of plant gene function and crop breeding. Moreover, we also discuss the improving points of genome editing in crop precision genetic improvement for further study.

Combinations of the Ghd7, Ghd8 and Hd1 genes largely define the ecogeographical adaptation and yield potential of cultivated rice.

Rice cultivars have been adapted to favorable ecological regions and cropping seasons. Although several heading date genes have separately made contributions to this adaptation, the roles of gene combinations are still unclear. We employed a map-based cloning approach to isolate a heading date gene, which coordinated the interaction between Ghd7 and Ghd8 to greatly delay rice heading. We resequenced these three genes in a germplasm collection to analyze natural variation. Map-based cloning demonstrated that the gene largely affecting the interaction between Ghd7 and Ghd8 was Hd1. Natural variation analysis showed that a combination of loss-of-function alleles of Ghd7, Ghd8 and Hd1 contributes to the expansion of rice cultivars to higher latitudes; by contrast, a combination of pre-existing strong alleles of Ghd7, Ghd8 and functional Hd1 (referred as SSF) is exclusively found where ancestral Asian cultivars originated. Other combinations have comparatively larger favorable ecological scopes and acceptable grain yield. Our results indicate that the combinations of Ghd7, Ghd8 and Hd1 largely define the ecogeographical adaptation and yield potential in rice cultivars. Breeding varieties with the SSF combination are recommended for tropical regions to fully utilize available energy and light resources and thus produce greater yields.

Natural variation and artificial selection in four genes determine grain shape in rice.

The size of cultivated rice (Oryza sativa) grains has been altered by both domestication and artificial selection over the course of evolutionary history. Several quantitative trait loci (QTLs) for grain size have been cloned in the past 10 yr. To explore the natural variation in these QTLs, resequencing of grain width and weight 2 (GW2), grain size 5 (GS5) and QTL for seed width 5 (qSW5) and genotyping of grain size 3 (GS3) were performed in the germplasms of 127 varieties of rice (O. sativa) and 10-15 samples of wild rice (Oryza rufipogon). Ten, 10 and 15 haplotypes were observed for GW2, GS5 and qSW5. qSW5 and GS3 had the strongest effects on grain size, which have been widely utilized in rice production, whereas GW2 and GS5 showed more modest effects. GS5 showed small sequence variations in O. sativa germplasm and that of its progenitor O. rufipogon. qSW5 exhibited the highest level of nucleotide diversity. GW2 showed signs of purifying selection. The four grain size genes experienced different selection intensities depending on their genetic effects. In the indica population, linkage disequilibrium (LD) was detected among GS3, qSW5 and GS5. The substantial genetic variation in these four genes provides the flexibility needed to design various rice grain shapes. These findings provide insight into the evolutionary features of grain size genes in rice.

Identification of genomic regions controlling adult-plant stripe rust resistance in Chinese landrace Pingyuan 50 through bulked segregant analysis.

ABSTRACT Stripe rust, caused by Puccinia striiformis f. sp. tritici, is one of the most widespread and destructive wheat diseases worldwide. Growing resistant cultivars with adult-plant resistance (APR) is an effective approach for the control of the disease. In this study, 540 simple sequence repeat markers were screened to map quantitative trait loci (QTL) for APR to stripe rust in a doubled haploid (DH) population of 137 lines derived from the cross Pingyuan 50 x Mingxian 169. The DH lines were planted in randomized complete blocks with three replicates in Gansu and Sichuan provinces during the 2005-06, 2006-07, and 2007-08 cropping seasons, providing data for four environments. Artificial inoculations were carried out in Gansu and Sichuan with the prevalent Chinese race CYR32. Broad-sense heritability of resistance to stripe rust for maximum disease severity was 0.91, based on the mean value averaged across four environments. Inclusive composite interval mapping detected three QTL for APR to stripe rust on chromosomes 2BS, 5AL, and 6BS, designated QYr.caas-2BS, QYr.caas-5AL, and QYr.caas-6BS, respectively, separately explaining from 4.5 to 19.9% of the phenotypic variation. QYr.caas-5AL, different from QTL previously reported, was flanked by microsatellite markers Xwmc410 and Xbarc261, and accounted for 5.0 to 19.9% of phenotypic variance. Molecular markers closely linked to the QTL could be used in marker-assisted selection for APR to stripe rust in wheat breeding programs.

QTL mapping for adult-plant resistance to stripe rust in Italian common wheat cultivars Libellula and Strampelli.

Italian common wheat cultivars Libellula and Strampelli, grown for over three decades in Gansu province of China, have shown effective resistance to stripe rust. To elucidate the genetic basis of the resistance, F(3) populations were developed from crosses between the two cultivars and susceptible Chinese wheat cultivar Huixianhong. The F(3) lines were evaluated for disease severity in Beijing, Gansu and Sichuan from 2005 to 2008. Joint- and single-environment analyses by composite interval mapping identified five quantitative trait loci (QTLs) in Libellula for reduced stripe rust severity, designated QYr.caas-2DS, QYr.caas-4BL, QYr.caas-5BL.1, QYr.caas-5BL.2 and QYr.caas-7DS, and explained 8.1-12.4, 3.6-5.1, 3.4-8.6, 2.6 and 14.6-35.0%, respectively, of the phenotypic variance across four environments. Six interactions between different pairs of QTLs explained 3.2-7.1% of the phenotypic variance. The QTLs QYr.caas-4BL, QYr.caas-5BL.1 and QYr.caas-7DS were also detected in Strampelli, explaining 4.5, 2.9-5.5 and 17.1-39.1% of phenotypic variance, respectively, across five environments. Three interactions between different pairs of QTLs accounted for 6.1-35.0% of the phenotypic variance. The QTL QYr.caas-7DS flanked by markers csLV34 and Xgwm295 showed the largest effect for resistance to stripe rust. Sequence analyses confirmed that the lines with the QYr.caas-7DS allele for resistance carried the resistance allele of the Yr18/Lr34 gene. Our results indicated that the adult-plant resistance gene Yr18 and several minor genes confer effective durable resistance to stripe rust in Libellula and Strampelli.