J-family genes redundantly regulate flowering time and increase yield in soybean

Soybean(Glycine max)is a short-day crop whose flowering time is regulated by photoperiod.The longjuvenile trait extends its vegetative phase and increases yield under short-day conditions.Natural variation in J,the major locus controlling this trait,modulates flowering time.We report that the three J-family genes influence soybean flowering time,with the triple mutant Guangzhou Mammoth-2 flowering late under short days by inhibiting transcription of E1-family genes.J-family genes offer promising allelic combinations for breeding.

Molecular mechanism of flowering time regulation in Brassica rapa:similarities and differences with Arabidopsis

Properly regulated flowering time is pivotal for successful plant reproduction.The floral transition from vegetative growth to reproductive growth is regulated by a complex gene regulatory network that integrates environmental signals and internal conditions to ensure that flowering takes place under favorable conditions.Brassica rapa is a diploid Cruciferae species that includes several varieties that are cultivated as vegetable or oil crops.Flowering time is one of the most important agricultural traits of B.rapa crops because of its influence on yield and quality.The transition to flowering in B.rapa is regulated by several environmental and developmental cues,which are perceived by several signaling pathways,including the vernalization pathway,the autonomous pathway,the circadian clock,the thermosensory pathway,and gibberellin(GA)signaling.These signals are integrated to control the expression of floral integrators BrFTs and BrSOC1s to regulate flowering.In this review,we summarized current research advances on the molecular mechanisms that govern flowering time regulation in B.rapa and compare this to what is known in Arabidopsis.

GmAP1d regulates flowering time under long-day photoperiods in soybean

Flowering time is important for adaptation of soybean(Glycine max)to different environments.Here,we conducted a genome-wide association study of flowering time using a panel of 1490 cultivated soybean accessions.We identified three strong signals at the qFT02-2 locus(Chr02:12037319–12238569),which were associated with flowering time in three environments:Gongzhuling,Mengcheng,and Nanchang.By analyzing linkage disequilibrium,gene expression patterns,gene annotation,and the diversity of variants,we identified an AP1 homolog as the candidate gene for the qFT02-2 locus,which we named GmAP1d.Only one nonsynonymous polymorphism existed among 1490 soybean accessions at position Chr02:12087053.Accessions carrying the Chr02:12087053-T allele flowered significantly earlier than those carrying the Chr02:12087053-A allele.Thus,we developed a cleaved amplified polymorphic sequence(CAPS)marker for the SNP at Chr02:12087053,which is suitable for marker-assisted breeding of flowering time.Knockout of GmAP1d in the‘Williams 82’background by gene editing promoted flowering under long-day conditions,confirming that GmAP1d is the causal gene for qFT02-2.An analysis of the region surrounding GmAP1d revealed that GmAP1d was artificially selected during the genetic improvement of soybean.Through stepwise selection,the proportion of modern cultivars carrying the Chr02:12087053-T allele has increased,and this allele has become nearly fixed(95%)in northern China.These findings provide a theoretical basis for better understanding the molecular regulatory mechanism of flowering time in soybean and a target gene that can be used for breeding modern soybean cultivars adapted to different latitudes.

Two soybean homologues of TERMINAL FLOWER 1 control flowering time under long day conditions

Flowering time is a key agronomic trait that directly affect the adaptation and yield of soybean.After whole genome duplications,about 75%of genes being represented by multiple copies in soybean.There are four TERMINAL FLOWER 1(TFL1)genes in soybean,and the TFL1b(Dt1)has been characterized as the determinant of stem growth habit.The function of other TFL1 homologs in soybean is still unclear.Here,we generated knockout mutants by CRISPR/Cas9 genome editing technology and found that the tfl1c/tfl1d double mutants flowered significantly earlier than wild-type plants.We investigated that TFL1c and TFL1d could physically interact with the b ZIP transcription factor FDc1 and bind to the promoter of APETALA1a(AP1a).RNA-seq and q RT-PCR analyses indicated that TFL1c and TFL1d repressed the expressions of the four AP1 homologs and delayed the flowering time in soybean.The two genes play important roles in the regulation of flowering time in soybean and mainly act as the flowering inhibitors under long-day conditions.Our results identify novel components in the flowering-time regulation network of soybean and will be invaluable for molecular breeding of improved soybean yield.

A differentially methylated region of the ZmCCT10 promoter affects flowering time in hybrid maize

Flowering time(FT) is a key maize domestication trait, variation in which allows maize to grow in a wide range of latitudes. Although previous studies have investigated the genetic control of FT-related traits per se, few studies of FT hybrid performance have been published. We characterized the genomic architecture associated with hybrid performance for FT in a hybrid panel by testcrossing Chang 7–2 with 328Ye478 × Qi319 recombinant inbred lines(RILs). We identified 11 quantitative trait loci(QTL) for hybrid performance in FT-related traits, including a major QTL qFH10 that controls hybrid performance and heterosis in a summer maize-growing region. However, this locus acts in regulating FT traits per se only in a spring maize-growing region. We validated ZmCCT10 as a candidate gene for qFH10 and found that differences between hybrids and their parental lines in DNA methylation in the differentially methylated region(DMR, –700 to –1520) of the ZmCCT10 promoter affected gene expression pattern and thereby FT in the summer maize-growing region.

QTL effects and epistatic interaction for flowering time and branch number in a soybean mapping population of Japanese×Chinese cultivars

Flowering time and branching type are important agronomic traits related to the adaptability and yield of soybean. Molecular bases for major flowering time or maturity loci, E1 to E4, have been identified. However, more flowering time genes in cultivars with different genetic backgrounds are needed to be mapped and cloned for a better understanding of flowering time regulation in soybean. In this study, we developed a population of Japanese cultivar（Toyomusume）×Chinese cultivar（Suinong 10） to map novel quantitative trait locus（QTL） for flowering time and branch number. A genetic linkage map of a F_2 population was constructed using 1 306 polymorphic single nucleotide polymorphism（SNP） markers using Illumina Soy SNP8 ki Select Bead Chip containing 7 189（SNPs）. Two major QTLs at E1 and E9, and two minor QTLs at a novel locus, qFT2_1 and at E3 region were mapped. Using other sets of F_2 populations and their derived progenies, the existence of a novel QTL of qFT2_1 was verified. qBR6_1, the major QTL for branch number was mapped to the proximate to the E1 gene, inferring that E1 gene or neighboring genetic factor is significantly contributing to the branch number.

Co-location of QTL for Sclerotinia stem rot resistance and flowering time in Brassica napus

Sclerotinia stem rot(SSR) caused by Sclerotinia sclerotiorum(Lib.) de Bary is one of the most devastating diseases of Brassica napus worldwide. Both SSR resistance and flowering time(FT) adaptation are major breeding goals in B. napus. However, early maturing rapeseed varieties, which are important for rice-rapeseed rotation in China, are often highly susceptible to SSR. Here, we found that SSR resistance was significantly negatively correlated with FT in a natural population containing 521 rapeseed inbred lines and a double haploid(DH) population with 150 individual lines, both of which had great variation in FT. Four chromosomal regions on A2, A6, C2, and C8 affecting both SSR resistance and FT were identified using quantitative trait loci(QTL) mapping after constructing a high-density genetic map based on single nucleotide polymorphism markers in the DH population.Furthermore, we aligned QTL for the two traits identified in the present and previous studies to the B. napus reference genome, and identified four colocalized QTL hotspots of SSR resistance and FT on A2(0–7.7 Mb), A3(0.8–7.5 Mb), C2(0–15.2 Mb), and C6(20.2–36.6 Mb). Our results revealed a genetic link between SSR resistance and FT in B.napus, which should facilitate the development of effective strategies in both early maturing and SSR resistance breeding and in map-based cloning of SSR resistance QTL.

矮牵牛DFR-A基因的克隆与表达分析

通过分离矮牵牛DFR-A基因,研究其在不同部位的表达规律和酶活性变化,为花色素苷合成的结构基因分离及其他研究奠定基础。基于已公布的矮牵牛DFR序列,直接从矮牵牛花瓣中分离了DFRA基因序列,利用实时荧光定量PCR技术研究其在不同器官中的表达特性,同时测定了不同器官中的DFR酶活性。(1)矮牵牛DFR-A基因cDNA序列为1473 bp,该基因最大开放阅读框为1143 bp,编码380个氨基酸。(2)DFR-A基因在矮牵牛各种器官中均有表达,但不同组织中表达量存在差异,在花药中的表达量最高,其次是初开和盛开花瓣中,各期花蕾中的表达量都较低,而在叶中表达量最低。(3)DFR酶在叶片及花药中几乎无活性。在初开花瓣中达到最高峰。(4)除花药以外的部位,DFR酶活性与DFR mRNA浓度存在线性相关性。矮牵牛DFR酶活性主要受转录调控,活性在初开花瓣中最高。

Accession-specific flowering time variation in response to nitrate fluctuation in Arabidopsis thaliana

Flowering time,a key transition point from vegetative to reproductive growth,is regulated by an intrinsic complex of endogenous and exogenous signals including nutrient status.For hundreds of years,nitrogen has been well known to modulate flowering time,but the molecular genetic basis on how plants adapt to ever-changing nitrogen availability remains not fully explored.Here we explore how Arabidopsis natural variation in flowering time responds to nitrate fluctuation.Upon nitrate availability change,we detect accession-and photoperiod-specific flowering responses,which also feature a accession-specific dependency on growth traits.The flowering time variation correlates well with the expression of floral integrators,SOC1 and FT,in an accession-specific manner.We find that gene expression variation of key hub genes in the photoperiod-circadian-clock(GI),aging(SPLs)and autonomous(FLC)pathways associates with the expression change of these integrators,hence flowering time variation.Our results thus shed light on the molecular genetic mechanisms on regulation of accession-and photoperiod-specific flowering time variation in response to nitrate availability.

Mapping QTL for flowering time-related traits under three plant densities in maize

Flowering time is an indicator of adaptation in maize and a key trait for selection in breeding.The genetic basis of flowering time in maize,especially in response to plant density,remains unclear.The objective of this study was to identify maize quantitative trait loci(QTL)associated with flowering time-related traits that are stably expressed under several plant densities and show additive effects that vary with plant density.Three hundred recombinant inbred lines(RIL)derived from a cross between Ye 478 and Qi 319,together with their parents,were planted at three plant densities(90,000,120,000,and 150,000 plants ha^(-1))in four environments.The five traits investigated were days to tasseling(DTT),days to silking(DTS),days to pollen shed(DTP),interval between anthesis and silking(ASI),and interval between tasseling and anthesis(TAI).A high-resolution bin map was used for QTL mapping.In the RIL population,the DTT,DTS,and DTP values increased with plant density,whereas the ASI and TAI values showed negligible response to plant density.A total of 72 QTL were identified for flowering time-related traits,including 15 stably expressed across environments.Maize flowering time under different densities seems to be regulated by complex pathways rather than by several major genes or an independent pathway.The effects of some stable QTL,especially qDTT8-1 and qDTT10-4,varied with plant density.Fine mapping and cloning of these QTL will shed light on the mechanism of flowering time and assist in breeding earlymaturing maize inbred lines and hybrids.

Development of mutants with varying flowering times by targeted editing of multiple SVP gene copies in Brassica napus L.

Manipulation of flowering time to develop cultivars with desired maturity dates is fundamental in plant breeding.It is desirable to generate polyploid rapeseed(Brassica napus L.)germplasm with varying flowering time controlled by a few genes.In the present study,Bna SVP,a rapeseed homolog of the Arabidopsis SVP(Short Vegetative Phase)gene,was characterized and a set of mutants was developed using a CRISPR/Cas9-based gene-editing tool.A single construct targeting multiple sites was successfully applied to precisely mutate four copies of Bna SVP.The induced mutations in these copies were stably transmitted to subsequent generations.Homozygous mutants with loss-of-function alleles and free transgenic elements were generated across the four Bna SVP homologs.All mutant T_(1)lines tested in two environments(summer and winter growing seasons)showed early-flowering phenotypes.The decrease in flowering time was correlated with the number of mutated Bna SVP alleles.The quadruple mutants showed the shortest flowering time,with a mean decrease of 40.6%–50.7%in length relative to the wild type under the two growth conditions.Our study demonstrates the quantitative involvement of Bna SVP copies in the regulation of flowering time and provides valuable resources for rapeseed breeding.

CRISPR/Cas9-engineered mutation to identify the roles of phytochromes in regulating photomorphogenesis and flowering time in soybean

Soybean(Glycine max)responds to ambient light variation by undergoing multiform morphological alterations,influencing its yield potential and stability in the field.Phytochromes(PHYs)are plant-specific red(R)and far-red(FR)light photoreceptors mediating photomorphogenesis and photoperiodic flowering.As an ancient tetraploid,soybean harbors four PHYA,two PHYB,and two PHYE paralogs.Except for GmPHYA2/E4 and GmPHYA3/E3,which have been identified as photoperiod-dependent flowering repressors,the functions of GmPHYs are still largely unclear.We generated a series of individual or combined mutations targeting the GmPHYA or GmPHYB genes using CRISPR/Cas9 technology.Phenotypic analysis revealed that GmPHYB1 mediates predominantly R-light induced photomorphogenesis,whereas GmPHYA2/E4 and GmPHYA3/E3,followed by GmPHYA1 and GmPHYB2,function redundantly and additively in mediating FR light responses in seedling stage.GmPHYA2/E4 and GmPHYA3/E3,with weak influence from GmPHYA1 and GmPHYA4,delay flowering time under natural long-day conditions.This study has demonstrated the diversified functions of GmPHYAs and GmPHYBs in regulating light response,and provides a core set of phytochrome mutant alleles for characterization of their functional mechanisms in regulating agronomic traits of soybean.

Multiplex CRISPR/Cas9-mediated knockout of soybean LNK2 advances flowering time

Flowering time is an important agronomic trait for soybean yield and adaptation. However, the genetic basis of soybean adaptation to diverse latitudes is still not clear. Four NIGHT LIGHT-INDUCIBLE AND CLOCK-REGULATED 2(LNK2) homeologs of Arabidopsis thaliana LNK2 were identified in soybean. Three single-guide RNAs were designed for editing the four LNK2 genes. A transgene-free homozygous quadruple mutant of the LNK2 genes was developed using the CRISPR(clustered regularly interspaced short palindromic repeats)/Cas9(CRISPR-associated protein 9). Under long-day(LD) conditions, the quadruple mutant flowered significantly earlier than the wild-type(WT). Quantitative real-time PCR(q RT-PCR)revealed that transcript levels of LNK2 were significantly lower in the quadruple mutant than in the WT under LD conditions. LNK2 promoted the expression of the legume-specific E1 gene and repressed the expression of FT2 a. Genetic markers were developed to identify LNK2 mutants for soybean breeding.These results indicate that CRISPR/Cas9-mediated targeted mutagenesis of four LNK2 genes shortens flowering time in soybean. Our findings identify novel components in flowering-time control in soybean and may be beneficial for further soybean breeding in high-latitude environments.

Expansion and expression diversity of FAR1/FRS-like genes provides insights into flowering time regulation in roses

Roses are important horticultural plants with enormous diversity in flowers and flowering behavior.However,molecular regulation of flowering time variation in roses remains poorly characterized.Here,we report an expansion of the FAR1/FRS-like genes that correlates well with the switch to prostrate-toerect growth of shoots upon flowering in Rosa wichuraiana‘Basye's Thornless'(BT).With the availability of the high-quality chromosome-level genome assembly for BT that we developed recently,we identified 91 RwFAR1/FRS-like genes,a significant expansion in contrast to 52 in Rosa chinensis‘Old Blush’(OB),a founder genotype in modern rose domestication.Rose FAR1/FRS-like proteins feature distinct variation in protein domain structures.The dispersed expansion of RwFAR1/FRS-like genes occurred specifically in clade I and II and is significantly associated with transposon insertion in BT.Most of the RwFAR1/FRS-like genes showed relatively higher expression level than their corresponding orthologs in OB.FAR1/FRS-like genes regulate light-signaling processes,shade avoidance,and flowering time in Arabidopsis thaliana.Therefore,the expansion and duplication of RwFAR1/FRS-like genes,followed by diversification in gene expression,might offer a novel leverage point for further understanding the molecular regulation of the variation in shoot-growth behavior and flowering time in roses.

Validation of SSR Markers Linked to Flowering Time QTLs in Sorghum through Progeny Test

Flowering time is critically important for crop yield, and detection of its genetic factors with strongly associated DNA markers is necessary in breeding programs. This study was undertaken to validate the quantitative trait loci （QTLs） underlying flowering time of sorghum based on the association between genotypes at SSR marker loci and flowering time in F3 family lines from self-pollinated heterozygous F2 plants developed by crossing between ＂SC112＂---an early flowering variety from Ethiopia and ＂Kikuchi Zairai＂--a late flowering variety from Japan. The results showed that the SSR markers linked to the QTLs on sorghum chromosomes 1, 2, 3, 5b, 7 and 8b were significantly （P 〈 0.05） associated with flowering time, and these markers and the QTLs reported previously are valid. On the other hand, the genotypes at the marker locus SB596 of qFT1-2 on chromosome 1 was not significantly associated with flowering time. The valid DNA markers, SB258 in qFTI-1, SB 1512 in qFT2, SB 1839 in qFT3, SB3369 in qFT5b, SB4096 in qFT7 and SB4540 and SB4660 in qFT8b, might be useful for DNA-marker assisted breeding.

茶树儿茶素合成酶基因DFR的表达特性

[目的]研究茶树儿茶素积累与二氢黄酮醇4-还原酶(dihydroflavonol 4-reductase, DFR)基因表达特性的关系。[方法]选取南昆山毛叶茶(Camellia ptilophylla Chang)和英红九号(Camellia sinensis Yinghong 9)茶树为材料,通过反转录PCR克隆茶树DFR基因,借助烟草瞬时表达技术对其表达蛋白进行亚细胞定位。经生物信息学分析,用特异性合成肽和匙孔血蓝蛋白偶联,通过免疫技术制备了该基因的特异性抗体。应用实时荧光定量PCR和Western blot技术测定DFR转录和翻译情况,并比较2种茶树中DFR基因在转录及翻译水平的差异。[结果]MY-DFR(南昆山毛叶茶DFR基因,Genbank登录号为MT875201)和YH-DFR(英红九号DFR基因,Genbank登录号为MT876617)间存在5个核苷酸碱基及相应5个编码氨基酸的差异,其表达蛋白亚细胞定位于细胞质;制备的DFR基因特异性兔源抗体效价为1∶80 000;DFR基因在南昆山毛叶茶中的转录水平极显著低于英红九号,但表达蛋白量显著高于英红九号。[结论]DFR基因在2种茶树中转录水平和翻译水平的差异体现了其表达的复杂性,推测茶树体内DFR基因的表达可能受到相关调控基因的调控。

The transcription factor HBF1 directly activates expression of multiple flowering time repressors to delay rice flowering

Flowering time(or heading date)is an important agronomic trait that determines the environmental adaptability and yield of many crops,including rice(Oryza sativa L.).Hd3a BINDING REPRESSOR FACTOR 1(HBF1),a basic leucine zipper transcription factor,delays flowering by decreasing the expression of Early heading date 1(Ehd1),Heading date 3a(Hd3a),and RICE FLOWERING LOCUS T 1(RFT1),but the underlying molecular mechanisms have not been fully elucidated.Here,we employed the hybrid transcriptional factor(HTF)strategy to enhance the transcriptional activity of HBF1 by fusing it to four copies of the activation domain from Herpes simplex virus VP16.We discovered that transgenic rice lines overexpressing HBF1-VP64(HBF1V)show significant delays in time to flower,compared to lines overexpressing HBF1-MYC or wild-type plants,via the Ehd1–Hd3a/RFT1 pathway,under both long-day and short-day conditions.Transcriptome deep sequencing analysis indicated that 19 WRKY family genes are upregulated in the HBF1V overexpression line.We demonstrate that the previously unknown gene,OsWRKY64,is a direct downstream target of HBF1 and represses flowering in rice,whereas three known flowering repressor genes,Days to heading 7(DTH7),CONSTANS 3(OsCO3),and OsWRKY104,are also direct target genes of HBF1 in flowering regulation.Taking these results together,we propose detailed molecular mechanisms by which HBF1 regulates the time to flower in rice.

GhAP1-D3 positively regulates flowering time and early maturity with no yield and fiber quality penalties in upland cotton

Flowering time(FTi)is a major factor determining how quickly cotton plants reach maturity.Early maturity greatly affects lint yield and fiber quality and is crucial for mechanical harvesting of cotton in northwestern China.Yet,few quantitative trait loci(QTLs)or genes regulating early maturity have been reported in cotton,and the underlying regulatory mechanisms are largely unknown.In this study,we characterized 152,68,and 101 loci that were significantly associated with the three key early maturity traits—FTi,flower and boll period(FBP)and whole growth period(WGP),respectively,via four genome-wide association study methods in upland cotton(Gossypium hirsutum).We focused on one major early maturity-related genomic region containing three single nucleotide polymorphisms on chromosome D03,and determined that GhAP1-D3,a gene homologous to Arabidopsis thaliana APETALA1(AP1),is the causal locus in this region.Transgenic plants overexpressing GhAP1-D3 showed significantly early flowering and early maturity without penalties for yield and fiber quality compared to wild-type(WT)plants.By contrast,the mutant lines of GhAP1-D3 generated by genome editing displayed markedly later flowering than the WT.GhAP1-D3 interacted with GhSOC1(SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1),a pivotal regulator of FTi,both in vitro and in vivo.Changes in GhAP1-D3 transcript levels clearly affected the expression of multiple key flowering regulatory genes.Additionally,DNA hypomethylation and high levels of H3K9ac affected strong expression of GhAP1-D3 in early-maturing cotton cultivars.We propose that epigenetic modifications modulate GhAP1-D3 expression to positively regulate FTi in cotton through interaction of the encoded GhAP1 with GhSOC1 and affecting the transcription of multiple flowering-related genes.These findings may also lay a foundation for breeding early-maturing cotton varieties in the future.

Picturing the Times

Girls with flowers on their heads at a celebration for the traditional Ngarot festival in West Java,Indonesia,on December 13,2023.The festival is a traditional ceremony to welcome the rice planting season.

Picturing the Times

Tourists sightsee on a boat in a massive lotus pond scenic area in Xinghua,Taizhou,east China’s Jiangsu Province,on August 5.When summer peaks,lotus flowers enter their blooming season,attracting throngs of tourists to admire their beauty.