Epigenetic control on transcription of vernalization genes and whole-genome gene expression profile induced by vernalization in common wheat

Vernalization is necessary for winter wheat to flower.However,it is unclear whether vernalization is also required for spring wheat,which is frequently sown in fall,and what molecular mechanisms underlie the vernalization response in wheat varieties.In this study,we examined the molecular mechanisms that regulate vernalization response in winter and spring wheat varieties.For this purpose,we determined how major vernalization genes(VRN1,VRN2,and VRN3)respond to vernalization in these varieties and whether modifications to histones play a role in changes in gene expression.We also identified genes that are differentially regulated in response to vernalization in winter and spring wheat varieties.We found that in winter wheat,but not in spring wheat,VRN1 expression decreases when returned to warm temperature following vernalization.This finding may be associated with differences between spring and winter wheat in the levels of tri-methylation of lysine 27 on histone H3(H3K27me3)and tri-methylation of lysine 4 on histone H3(H3K4me3)at the VRN1 gene.Analysis of winter wheat transcriptomes before and after vernalization revealed that vernalization influences the expression of several genes,including those involved in leucine catabolism,cysteine biosynthesis,and flavonoid biosynthesis.These findings provide new candidates for further study on the mechanism of vernalization regulation in wheat.

Border effects of the main and ratoon crops in the rice ratooning system

The border effect(BE)is widely observed in crop field experiments,and it has been extensively studied in many crops.However,only limited attention has been paid to the BE of ratoon rice.We conducted field experiments on ratoon rice in Qichun County,Hubei Province,Central China in 2018 and 2019 to compare the BE in the main and ratoon crops,and to quantify the contribution of BE in the main crop to that in the ratoon crop.The BE of two hybrid varieties was measured for the outermost,second outermost,and third outermost rows in each plot of both crops.To determine the contribution of BE between the two crops,portions of hills in the outermost and second outermost rows were uprooted during the harvest of the main crop so that the second and third outermost rows then became the outermost rows in the ratoon crop.Overall,the BE on grain yield was greater in the main crop than in the ratoon crop.In the main crop,the BE on grain yield was 98.3%in the outermost row,which was explained by the BE on panicles m^(–2),spikelets/panicle,spikelets m^(–2),and total dry weight.In the ratoon crop,the BE on grain yield was reduced to 60.9 and 27.6%with and without the contribution of the BE in the main crop,respectively.Consequently,55.1%of the BE on grain yield in the ratoon crop was contributed from the main crop.High stubble dry weight and non-structural carbohydrate(NSC)accumulation at the harvest of the main crop were responsible for the contribution of BE in the main crop to that in the ratoon crop.Our results suggest that increases in stubble dry weight and NSC accumulation at the harvest of the main crop could be important strategies for developing high-yielding cropping practices in the rice ratooning system.

Knock-out of GhPDCT with the CRISPR/Cas9 system increases the oleic acid content in cottonseed oil

Cotton is a pivotal economic crop for natural textile fibers that also serves as an important source of edible oil(Long et al.2023).Cottonseed oil contains approximately14%oleic acid and 59%linoleic acid.An increase in monounsaturated fatty acids,particularly oleic acid,enhances the oxidative stability and nutritional value of edible oil(Chen et al.2021).

Genetic analysis of GEFs and GDIs in rice reveals the roles of OsGEF5,OsGDI1,and OsGEF3 in the regulation of grain size and plant height

Guanine nucleotide exchange factors(GEFs)and guanine nucleotide-dissociation inhibitors(GDIs)regulate small GTPase proteins,which function as molecular switches in various signaling pathways,but their identification and functions in plants are not well understood.Using in-silico analysis and transgenic approaches,respectively,we dissected the evolutionary relationships and functions of all GEF and GDI genes in rice.Intron-exon distribution and phylogenetic analyses identified 30 GEF and 10 GDI genes in rice that shared close evolutionary relationships with other eukaryotes.Tissue-specific expression and co-expression analyses revealed that phylogenetically related genes had similar expression patterns.GEF and GDI genes were highly expressed in panicles,hulls,and stamens.Co-expression network analysis identified panicle and stamen-specific modules of co-expressed genes in both families.Mapping of these genes in known protein interactomes further identified two and one small G-protein sub-networks.A mutant library of GEF and GDI families was constructed by CRISPR knockout of each gene,and their genotypes and phenotypes were confirmed.Phenotype changes occurred with the mutation of only three genes(OsGEF5,OsGDI1,and OsGEF3).OsGEF5 and OsGDI1 single mutants exhibited significantly reduced height and longer and thinner grains,whereas OsGEF3 mutants had reduced grain length compared to the wild type.Haplotype and eGWAS analyses showed that natural variations in the three genes affected gene expression in reproductive tissues that were significantly associated with the phenotypic variation.BiFC assays demonstrated that GDI1 and GEF3 interacted with grain-size protein GS3,pointing to a role of these genes in the regulation of grain size and plant architecture connected to heterotrimeric G-proteins in rice.

Advancing approach and toolbox in optimization of chloroplast genetic transformation technology

Chloroplast is a discrete,highly structured,and semi-autonomous cellular organelle.The small genome of chloroplast makes it an up-and-coming platform for synthetic biology.As a special means of synthetic biology,chloroplast genetic engineering shows excellent potential in reconstructing various sophisticated metabolic pathways within the plants for specific purposes,such as improving crop photosynthetic capacity,enhancing plant stress resistance,and synthesizing new drugs and vaccines.However,many plant species exhibit limited efficiency or inability in chloroplast genetic transformation.Hence,new transformation technologies and tools are being constantly developed.In order to further expand and facilitate the application of chloroplast genetic engineering,this review summarizes the new technologies in chloroplast genetic transformation in recent years and discusses the choice of appropriate synthetic biological elements for the construction of efficient chloroplast transformation vectors.

Genetic Diversity,Population Structure,and Genome-Wide Association Study of Seven Agronomic Traits in 273 Diverse Upload Cotton Accessions

Upland cotton(Gossypium hirsutum)is the most important plant producing natural fibers for the textile industry.In this study,we first investigated the phenotypic variation of seven agronomic traits of 273 diverse cotton accessions in the years 2017 and 2018,which were from 18 geographical regions.We found large variations among the traits in different geographical regions and only half of the traits in either years 2017 or 2018 followed a normal distribution.We then genotyped the collection with 81,612 high quality SNPs.Phylogenetic tree and population structure revealed a diverse genetic structure of the core collection,and geographical diversification was an important factor,but account for part of the variances of genetic diversification.We then performed genome-wide association study for the seven traits in the years 2017 and 2018,and the average values of each trait in the two years,respectively.We identified a total of 19 significant marker-trait associations and found that Pollen Ole e 1 allergen/extension could be the candidate gene associated with the fall-off cotton bolls from the last three branches.In addition,large variations were observed for the heritability of traits in the years 2017 and 2018.These results provide new potential candidate genes for further functional validation,which could be useful for genetic improvement and breeding of new cotton cultivars with better agronomic performances.

Association mapping and domestication analysis to dissect genetic improvement process of upland cotton yield-related traits in China

Background: Cotton fiber yield is a complex trait,which can be influenced by multiple agronomic traits.Unravelling the genetic basis of cotton fiber yield-related traits contributes to genetic improvement of cotton.Results: In this study,503 upland cotton varieties covering the four breeding stages(BS1–BS4,1911–2011)in China were used for association mapping and domestication analysis.One hundred and forty SSR markers significantly associated with ten fiber yield-related traits were identified,among which,29 markers showed an increasing trend contribution to cotton yield-related traits from BS1 to BS4,and 26 markers showed decreased trend effect.Four favorable alleles of 9 major loci(R^(2)≥3)were strongly selected during the breeding stages,and the candidate genes of the four strongly selected alleles were predicated according to the gene function annotation and tissue expression data.Conclusions :The study not only uncovers the genetic basis of 10 cotton yield-related traits but also provides genetic evidence for cotton improvement during the cotton breeding process in China.

Genome-wide and candidate gene association studies identify BnPAP17 as conferring the utilization of organic phosphorus in oilseed rape

Phosphorus(P)is essential for living plants,and P deficiency is one of the key factors limiting the yield in rapeseed production worldwide.As the most important organ for plants,root morphology traits(RMTs)play a key role in P absorption.To investigate the genetic variability of RMT under low P availability,we dissected the genetic structure of RMTs by genome-wide association studies(GWAS),linkage mapping and candidate gene association studies(CGAS).A total of 52 suggestive loci were associated with RMTs under P stress conditions in 405 oilseed rape accessions.The purple acid phosphatase gene BnPAP17 was found to control the lateral root number(LRN)and root dry weight(RDW)under low P stress.The expression of BnPAP17 was increased in shoot tissue in P-efficient cultivars compared to root tissue and P-inefficient cultivars in response to low P stress.Moreover,the haplotype of BnPAP17^(Hap3)was detected for the selective breeding of P efficiency in oilseed rape.Over-expression of the BnPAP17^(Hap3)could promote the shoot and root growth with enhanced tolerance to low P stress and organic phosphorus(Po)utilization in oilseed rape.Collectively,these findings increase our understanding of the mechanisms underlying BnPAP17-mediated low P stress tolerance in oilseed rape.

Comprehensive integration of single-cell transcriptomic data illuminates the regulatory network architecture of plant cell fate specification

Plant morphogenesis relies on precise gene expression programs at the proper time and position which is orchestrated by transcription factors(TFs)in intricate regulatory networks in a cell-type specific manner.Here we introduced a comprehensive single-cell transcriptomic atlas of Arabidopsis seedlings.This atlas is the result of meticulous integration of 63 previously published scRNA-seq datasets,addressing batch effects and conserving biological variance.This integration spans a broad spectrum of tissues,including both below-and above-ground parts.Utilizing a rigorous approach for cell type annotation,we identified 47 distinct cell types or states,largely expanding our current view of plant cell compositions.We systematically constructed cell-type specific gene regulatory networks and uncovered key regulators that act in a coordinated manner to control cell-type specific gene expression.Taken together,our study not only offers extensive plant cell atlas exploration that serves as a valuable resource,but also provides molecular insights into gene-regulatory programs that varies from different cell types.

Identification of new cotton fiber-quality QTL by multiple genomic analyses and development of markers for genomic breeding

Cotton fiber is one of the main raw materials for the textile industry.In recent years,many cotton fiber quality QTL have been identified,but few were applied in breeding.In this study,a genome wide association study(GWAS)of fiber-quality traits in 265 upland cotton breeding intermediate lines(GhBreeding),combined with genome-wide selective sweep analysis(GSSA)and genomic selection(GS),revealed 25 QTL.Most of these QTL were ignored by only using GWAS.The CRISPR/Cas9 mutants of GhMYB_D13 had shorter fiber,which indicates the credibility of QTL to a certain extent.Then these QTL were verified in other cotton natural populations,5 stable QTL were found having broad potential for application in breeding.Additionally,among these 5 stable QTL,superior genotypes of 4 showed an enrichment in most improved new varieties widely cultivated currently.These findings provide insights for how to identify more QTL through combined multiple genomic analysis to apply in breeding.

EPSPS regulates cell elongation by disrupting the balance of lignin and flavonoid biosynthesis in cotton

EPSPS is a key gene in the shikimic acid synthesis pathway that has been widely used in breeding crops with herbicide resistance.However,its role in regulating cell elongation is poorly understood.Through the overexpression of EPSPS genes,we generated lines resistant to glyphosate that exhibit an unexpected dwarf phenotype.A representative line,DHR1,exhibits a stable dwarf phenotype throughout its entire growth period.Except for plant height,the other agronomic traits of DHR1 are similar to its transgenic explants ZM24.Paraffin section observations showed that DHR1 internodes are shortened due to reduced elongation and division of the internode cells.Exogenous hormones confirmed that DHR1 is not a classical brassinolide(BR)-or gibberellin(GA)-related dwarfing mutant.Hybridization analysis and fine mapping confirmed that the EPSPS gene is the causal gene for dwarfism,and the phenotype can be inherited in different genotypes.Transcriptome and metabolome analyses showed that genes associated with the phenylpropanoid synthesis pathway are enriched in DHR1 compared with ZM24.Flavonoid metabolites are enriched in DHR1,whereas lignin metabolites are reduced.The enhancement of flavonoids likely results in differential expression of auxin signal pathway genes and alters the auxin response,subsequently affecting cell elongation.This study provides a new strategy for generating dwarfs and will accelerate advancements in light simplification in the cultivation and mechanized harvesting of cotton.

The occurrence,inheritance,and segregation of complex genomic structural variation in synthetic Brassica napus

"Synthetic"allopolyploids recreated by interspecific hybridization play an important role in providing novel genomic variation for crop improvement.Such synthetic allopolyploids often undergo rapid genomic structural variation(SV).However,how such SV arises,is inherited and fixed,and how it affects important traits,has rarely been comprehensively and quantitively studied in advanced generation synthetic lines.A better understanding of these processes will aid breeders in knowing how to best utilize synthetic allopolyploids in breeding programs.Here,we analyzed three genetic mapping populations(735 DH lines)derived from crosses between advanced synthetic and conventional Brassica napus(rapeseed)lines,using whole-genome sequencing to determine genome composition.We observed high tolerance of large structural variants,particularly toward the telomeres,and preferential selection for balanced homoeologous exchanges(duplication/deletion events between the A and C genomes resulting in retention of gene/chromosome dosage between homoeologous chromosome pairs),including stable events involving whole chromosomes("pseudoeuploidy").Given the experimental design(all three populations shared a common parent),we were able to observe that parental SV was regularly inherited,showed genetic hitchhiking effects on segregation,and was one of the major factors inducing adjacent novel and larger SV.Surprisingly,novel SV occurred at low frequencies with no significant impacts on observed fertility and yield-related traits in the advanced generation synthetic lines.However,incorporating genome-wide SV in linkage mapping explained significantly more genetic variance for traits.Our results provide a framework for detecting and understanding the occurrence and inheritance of genomic SV in breeding programs,and support the use of synthetic parents as an important source of novel trait variation.

Development of a rapid and efficient system for CR genes identification based on hairy root transformation in Brassicaceae

Many economically important crops and vegetables belonging to the cruciferous family are heavily endangered by clubroot disease caused by Plasmodiophora brassicae infection.Breeding of clubroot resistant cultivars based on mapping and cloning of resistant genes is commonly regarded as the most cost-effective and efficient way to fight against this disease.The traditional way of R gene functional validation requires stable transformation that is both time-and labor-consuming.In this study,a rapid and efficient hairy-root transgenic protocol mediated by Agrobacterium rhizogenes was developed.The transformation positive rate was over 80%in Brassica napus showed by GUS reporter gene and this transformation only took 1/6 of the time compared with stable transformation.The system was applicable to different B.napus varieties and other cruciferous crops including Brassica rapa and Brassica oleracea.In particular,two known CR genes,CRA3.7.1 and CRA8.2.4 were used respectively,as example to show that the system works well for CR gene study combined with subsequent P.brassicae infection in B.napus.Most importantly,it works both in over-expression that led to disease resistance,as well as in RNAi which led to disease susceptible phenotype.Therefore,this system can be used in batch-wise identification of CR genes,and also offered the possibility of manipulating key genes within the P.brassicae genome that could improve our knowledge on host-pathogen interaction.

OsMYB84,a transcriptional regulator of OsCOPT2 and OsHMA5,modulates copper uptake and transport and yield production in rice

Transcription factors regulating crop uptake and translocation of the micronutrient Cu have not been identified.We isolated a novel R2R3-MYB transcription factor,OsMYB84,and showed that it was a positive regulator involved in uptake and transport of Cu via activation of OsCOPT2 and OsHMA expression.OsMYB84 was highly expressed in roots and anthers and induced by Cu.Overexpression of OsMYB84 promoted uptake and root-to-shoot translocation of Cu in rice,facilitated Cu distribution into grain and increased grain yield.In contrast,mutation of OsMYB84 reduced Cu concentration in xylem sap.OsMYB84 bound to the promoter region of OsCOPT2 and OsHMA5 and upregulated their expression.OsCOPT2 mutants showed reduced uptake of Cu and OsHMA5 overexpression lines showed increased root-to-shoot translocation of Cu.

How rice organs are colored: The genetic basis of anthocyanin biosynthesis in rice

Anthocyanins are a major subclass of flavonoids that have diverse biological functions and benefit human health.In rice(Oryza sativa),the various colors shown by organs are due mainly to the accumulation of anthocyanins and are traits associated with domestication.Elucidating the genetic basis of anthocyanin biosynthesis in rice would support the engineering of anthocyanins as well as shedding light on the evolutionary history of O.sativa.We summarize recent progress in rice anthocyanin biosynthesis research,including gene cloning,biosynthetic pathway discovery,and study of the domestication process.We discuss the application of anthocyanin biosynthesis genes in rice breeding.Our object is to broaden knowledge of the genetic basis of anthocyanin biosynthesis in rice and support the breeding of novel rice cultivars.

Heavy soil drying during mid-to-late grain filling stage of the main crop to reduce yield loss of the ratoon crop in a mechanized rice ratooning system

Yield loss(Y_(Loss)) in the ratoon crop due to crushing damage to left stubble from mechanical harvesting of the main crop is a constraint for wide adoption of mechanized rice ratooning technology.Soil drying before the harvest of the main crop has been proposed to overcome this problem.The objective of this study was to determine the effect of soil drying during the mid-to-late grain filling stage of the main crop on grain yield of the ratoon crop in a mechanized rice ratooning system.Field experiments were conducted to compare Y_(Loss) between light(LD) and heavy(HD) soil drying treatments in Hubei province,central China in 2017 and 2018.Y_(Loss) was calculated as the percentage of yield reduction in the ratoon crop with the main crop harvested mechanically,relative to the grain yield of the ratoon crop with the main crop harvested manually.In comparison with LD,soil hardness was increased by 42.8%-84.7% in HD at the 5-20 cm soil depth at maturity of the main crop.Soil hardness at 5 and 10 cm depths reached respectively 4.05 and 7.07 kg cm^(-2) in HD.Soil drying treatment did not significantly affect the grain yield of the main crop.Under mechanical harvesting of the main crop,HD increased the grain yield of the ratoon crop by 9.4% relative to LD.Consequently,Y_(Loss) was only 3.4% in HD,in contrast to 16.3% in LD.The differences in grain yield and Y_(Loos) between the two soil drying treatments were explained mainly by panicles m^(-2),which was increased significantly by HD in the track zone of the ratoon crop compared with LD.These results suggest that heavy soil drying practice during the mid-to-late grain filling stage of the main crop is effective for reducing Y_(Loss) of the ratoon crop in a mechanized rice ratooning system.

Conversion of the Statistical Combining Ability into a Genetic Concept

Since the combining ability was proposed in 1942, efforts to uncover the genetic basis underlying this phenomenon have been ongoing for nearly 70 yr, with little success. Some breeding strategies based on evaluation of combining ability have been produced, and are still extensively used in hybrid breeding. In this review, the genetic basis underlying these breeding strategies is discussed, and a potential genetic control of general combining ability （GCA） is postulated. We suggested that GCA and the yields of inbred lines might be genetically controlled by different sets of loci on the maize genome that are transmitted into offspring. Different inbred lines might possess different favorable alleles for GCA. In hybrids, loci involved in multiple pathways, which are directly or indirectly associated with yield performance, might be regulated by GCA loci. In addition, a case of GCA mapping using a set of testcross progeny from introgression lines is provided.

Effects of nitrogen management on the ratoon crop yield and head rice yield in South USA

Ratoon rice cropping is an important component of the rice cropping system in Texas and south Louisiana,USA,and expanded to Asian countries in 1970.Two field studies were conducted with widely planted rice(Oryza sativa L.)cultivars at Eagle Lake,Texas,USA to determine the effects of nitrogen(N)management in main(first)crop(MC)and ratoon(second)crop(RC)on RC yield.In 2012 and 2013,one cultivar(Presidio)was adopted to determine the effects of RC N management on ratoon yield and head rice yield.In 2016 and 2017,CL153,CL163 and CL272 in addition to Presidio were adopted to examine the effect of MC N management on ratoon yield and head rice yield.N applied at preflood after MC harvest considerably improved RC yield.Application of 99 kg N ha^(–1)at preflood after MC harvest was practically adequate for RC regrowth,development and approaching the yield potential for Presidio.RC could produce quite high average grain yields of 5.90 to 6.53 t ha–1 in 2012 and 2013,respectively.Main crop N rate only significantly affected MC yield;however,given N applied of 99 kg ha^(–1)at preflood after MC harvest,ratoon yield was not significantly affected by MC N rate.Neither the main nor ratoon crop N management had a significant effect on RC head rice yield.Considerable RC head rice yields(55–65%)were observed in all of the four cultivars and 4 years except for CL272 in 2016.These results indicat that without very high N fertilizer application,rice ratoon crop could produce a considerable grain yield and an expectative head rice yield.Rice ratooning could be a practical way to increase rice yields with the minimal input in south Texas and regions with a similar climate.

Mapping and genetic validation of a grain size QTL qGS7.1 in rice(Oryza sativa L.)

Grain size is a major determinant of grain weight, which is one of the components of rice yield. The objective o this study was to identify novel, and important quantitative trait loci(QTLs) for grain size and weight in rice. QTLs were mapped using a BC4F4 population including 192 backcross inbred lines(BILs) derived from a backcross between Xiaolijing(XLJ) and recombinant inbred lines(RILs). The mapping population was planted in both Lingshui(Hainan, 2015) and Fuyang(Zhejiang, 2016), with the short-and long-day conditions, respectively. A total of 10 QTLs for grain length, four for grain width, four for the ratio of grain length to width, and 11 for grain weight were detected in at least one environment and were distributed across 11 chromosomes. The phenotypic variance explained ranged from 6.76–25.68%, 14.30–34.03%, 5.28–26.50%, and 3.01–22.87% for grain length, grain width, the ratio of grain length to width, and thousand grain weight, respectively. Using the sequential residual heterozygotes(SeqRHs) method, qGS7.1, a QTL for grain size and weight, was mapped in a 3.2-Mb interval on chromosome 7. No QTLs about grain size and weight were reported in previous studies in this region, providing a good candidate for functional analysis and breeding utilization.

Improvements of TKC Technology Accelerate Isolation of Transgene-Free CRISPR/Cas9-Edited Rice Plants

Elimination of the CRISPR/Cas9 constructs in edited plants is a prerequisite for assessing genetic stability, conducting phenotypic characterization, and applying for commercialization of the plants. However, removal of the CRISPR/Cas9 transgenes by genetic segregation and by backcross is laborious and time consuming. We previously reported the development of the transgene killer CRISPR(TKC) technology that uses a pair of suicide genes to trigger self-elimination of the transgenes without compromising gene editing efficiency. The TKC technology enables isolation of transgene-free CRISPR-edited plants within a single generation, greatly accelerating crop improvements. Here, we presented two new TKC vectors that show great efficiency in both editing the target gene and in undergoing self-elimination of the transgenes. The new vectors replaced the CaMV35 S promoter used in our previous TKC vector with two rice promoters to drive one of the suicide genes, providing advantages over our previous TKC vector under certain conditions. The vectors reported here offered more options and flexibility to conduct gene editing experiments in rice.