Investigation of Aegilops umbellulata for stripe rust resistance,heading date,and the contents of iron,zinc,and gluten protein

Aegilops umbellulata(UU)is a wheat wild relative that has potential use in the genetic improvement of wheat.In this study,46 Ae.umbellulata accessions were investigated for stripe rust resistance,heading date(HD),and the contents of iron(Fe),zinc(Zn),and seed gluten proteins.Forty-two of the accessions were classified as resistant to stripe rust,while the other four accessions were classified as susceptible to stripe rust in four environments.The average HD of Ae.umbellulata was significantly longer than that of three common wheat cultivars(180.9 d vs.137.0 d),with the exception of PI226500(138.9 d).The Ae.umbellulata accessions also showed high variability in Fe(69.74-348.09 mg kg^(-1))and Zn(49.83-101.65 mg kg^(-1))contents.Three accessions(viz.,PI542362,PI542363,and PI554399)showed relatively higher Fe(230.96-348.09 mg kg^(-1))and Zn(92.46-101.65 mg kg^(-1))contents than the others.The Fe content of Ae.umbellulata was similar to those of Ae.comosa and Ae.markgrafii but higher than those of Ae.tauschii and common wheat.Aegilops umbellulata showed a higher Zn content than Ae.tauschii,Ae.comosa,and common wheat,but a lower content than Ae.markgrafii.Furthermore,Ae.umbellulata had the highest proportion of γ-gliadin among all the species investigated(Ae.umbellulata vs.other species=mean 72.11%vs.49.37%;range:55.33-86.99%vs.29.60-67.91%).These results demonstrated that Ae.umbellulata exhibits great diversity in the investigated traits,so it can provide a potential gene pool for the genetic improvement of these traits in wheat.

Avoidance of Linkage Drag Between Blast Resistance Gene and the QTL Conditioning Spikelet Fertility Based on Genotype Selection Against Heading Date in Rice

Previous study showed that a linkage drag between a blast resistance gene Pi25（t） and QTLs conditioning spikelet fertility （qSF-6） and number of filled grains per panicle （qNFGP-6） was detected on the short arm of chromosome 6. A larger population was used for further verification, and the results confirmed the linkage drag between the blast resistance gene and QTL conditioning spikelet fertility, other than QTL conditioning number of filled grains per panicle. Breakdown or avoidance of the linkage drag could be achieved by selection against the genotype background of a heading-date gene （qHD-7） that resided in the region between RM2 and RM214 on chromosome 7. For further validation, two lines with almost identical genotypes on all chromosomal regions except the Pi25（t） region on chromosome 6 were chosen to develop a new population The results showed that qSF-6 could be further subdivided into qSF-6-1 and qSF-6-2. When the genotype of the region between RM2 and RM214 was from rice variety Zhong 156, the linkage drag between Pi25（t） and qSF-6-2 was detected and the allele of qSF-6-2 from rice variety Gumei 2 reduced the spikelet fertility. When the genotype of the region between RM2 and RM214 was from Gumei 2, no linkage drag was detected. This indicates that the linkage drag between the blast resistance gene and the QTL conditioning spikelet fertility could be broken down or avoided under a certain background genotype selection against heading-date and provides a marker aided solution for high level of blast resistance and yield breeding in rice and other crops as well.

Characterization of the Main Effects, Epistatic Effects and Their Environmental Interactions of QTL on the Genetic Basis of Plant Height and Heading Date in Rice

Main-effect QTL, epistatic effects and their interactions with environment are important genetic components of quantitativetraits. In this study, we analyzed the QTL, epistatic effects and QTL by environment interactions (QE) underlying plantheight and heading date, using a doubled-haploid ( DH) population consisting of 190 lines from the cross between anindica parent Zhenshan 97 and a japonica parent Wuyujing 2, and tested in two-year replicated field trials. A geneticlinkage map with 179 SSR (simple sequence repeat) marker loci was constructed. A mixed linear model approach wasapplied to detect QTL, digenic interactions and QEs for the two traits. In total, 20 main-effect QTLs, 9 digenic interactionsinvolving 18 loci, and 5 QTL by environment interactions were found to be responsible for the two traits. No interactionswere detected between the digenic interaction and environment. The amounts of variations explained by QTLs of maineffect were 53.9% for plant height and 57.8% for heading date, larger than that explained by epistasis and QEs. However,the epistasis and QE interactions sometimes accounted for a significant part of phenotypic variation and should not bedisregarded.

Quantitative Trait Loci for Heading Date and Their Relationship with Genetic Control of Yield Traits in Rice (Oryza sativa)

Grain yield and heading date are key factors determining the commercial potential of a rice variety. Mapping of quantitative trait loci （QTLs） in rice has been advanced from primary mapping to gene cloning, and heading date and yield traits have always attracted the greatest attention. In this review, genomic distribution of QTLs for heading date detected in populations derived from intra-specific crosses of Asian cultivated rice （Oryza sativa） was summarized, and their relationship with the genetic control of yield traits was analyzed. The information could be useful in the identification of QTLs for heading date and yield traits that are promising for the improvement of rice varieties.

TaGW2L,a GW2-like RING finger E3 ligase,positively regulates heading date in common wheat(Triticum aestivum L.)

RING finger E3 ligases play an important role in regulating plant growth and development by mediating substrate degradation.In this study,we identified TaGW2L,encoding a Grain width and weight2(GW2)-like RING finger E3 ligase,as a novel positive regulator of heading date in wheat(Triticum aestivum L.).TaGW2L exhibited high amino acid sequence similarities with TaGW2 homoeologs,particularly in the conserved RING finger domain.Expression analysis indicated that TaGW2L was constitutively expressed in various wheat tissues.TaGW2L showed transactivation activity in yeast and could interact with the ubiquitin-conjugating enzymes E2_(s).An in vitro ubiquitination assay verified that TaGW2L possessed a similar E3 ligase activity to TaGW2.Overexpression of the TaGW2L-7A homoeolog in wheat led to a significantly earlier heading date under both natural conditions and long-day conditions.Transcriptome analysis revealed that multiple known genes positively regulating wheat heading were significantly upregulated in the TaGW2L-7A-overexpression plants compared with the wild-type control.Together,our findings shed light on the role of TaGW2L in wheat heading date and provide potential applications of TaGW2L for the adaptation improvement of crops.

Identification and Fine Mapping of Heading Date Related Mutant Gene in Rice

A rice heading-date-related mutant was isolated from a ^60Co-y-ray-induced mutation pool of Zhejing 22, a conventional japonica cultivar in Zhejiang Province, China. The mutant was characterized by a delayed heading date of almost 20 d longer than the wild type plant. Genetic analysis revealed that the mutation was controlled by a single nuclear-encoded recessive gene that was designed as HD（t） （heading date tentatively）. To isolate the HD（t） gene, a map-based cloning approach was employed using 479 F2 mutant individual plants derived from the cross between the hd（t） mutant （japonica） x Zhenshan 97 （indica）. Finally, the HD（t） gene was mapped to an approximate 53 kb region between the insertion and deletion （InDel） markers of 10-61W and 10-66W on chromosome 10. According to the genome sequence of Nipponbare, the target region contains 11 annotated genes. It is helpful for future cloning of HD（t） gene based on this fine mapping results.

NRL3 Interacts with OsK4 to Regulate Heading Date in Rice

NRL3 is essential for the growth and development of rice leaves.In this study,we found that the loss function of NRL3 also delayed heading date under natural long daylight and short daylight conditions.The yeast two-hybrid and the bimolecular fluorescence complementation proved that NRL3interacts with OsK4,a Snf1-related kinase.OsK4 localized to the nucleus and expressed in various rice tissues.The rhythmic expression pattern of Os K4 was similar to NRL3 under long daylight and short daylight conditions.Knock-out mutants of Os K4 exhibited early heading under long daylight conditions,indicating that it acts as a negative regulator of heading date in rice.Interestingly,the OsK4 mutant under the nrl3 mutant background rescued the late heading phenotype of nrl3 under long daylight conditions,suggesting that Os K4 functions downstream of NRL3.Moreover,both NRL3 and Os K4 controlled heading date through regulating the expression of Hd3a and RFT1 genes.These findings shed light on the heading date regulation in rice and provide a sound theoretical base to improve regional adaptability of rice.

Clock component OsPRR59 delays heading date by repressing transcription of Ehd3 in rice

Heading date(or flowering time),an important agronomic trait in crop species,is closely associated with regional adaptation and yield.Members of the Pseudo-Response Regulator(PRR)family play key roles in regulating flowering.However,their role and molecular mechanism controlling heading date in rice is not very clear.Here,we identified rice OsPRR protein,OsPRR59,which delayed heading under longday conditions.OsPRR59 positively regulates yield by affecting plant height,secondary branches number per panicle,grain number per panicle,seed setting rate,and grain weight per plant.OsPRR59 is expressed in most tissues and its protein is localized to the nucleus.We also found that OsPRR59 directly binds to the promoter of Ehd3 to inhibit its expression.Compared with the WT,osprr59 ehd3 showed a significantly delayed heading phenotype,as did the ehd3 mutant.This was opposite to the phenotype of the osprr59 mutant,confirming that Ehd3 acted downstream of OsPRR59 in regulating rice flowering.Our results identified a direct regulator of Ehd3,and revealed a novel molecular mechanism of clock component OsPRR proteins in regulating heading date and provide a new genetic resource for fine-tuning heading date in rice.

Mapping of the heading date gene HdAey2280 in Aegilops tauschii

An optimum heading date is essential for sustainable crop productivity and ensuring high yields. In the present study, F2：3 populations were generated by crossing an early-heading accession, Y2280, with a late-heading accession, Y2282. The heading dates of the F2 and F3 populations were investigated in a field study. Using publicly available simple sequence repeat （SSR） markers, the early heading date gene HdAey2280 was mapped onto Aegilops tauschfi chromosome 7DS between the flanking markers wmc438 and barc126 at distances of 15 and 9.1 cM, respectively. Further analysis indicated that HdAey2280 is a novel heading date gene. New SSR markers were developed based on the Ae. tauschfi draft genome sequence, resulting in four new markers that were linked to the heading date gene HdAey2280. The closest distance of these markers was 1.9 cM away from the gene. The results collected in this study will serve as a framework for map-based cloning and marker-assisted selection in wheat breeding programs in the future.

QTL Detection and Epistasis Analysis for Heading Date Using Single Segment Substitution Lines in Rice(Oryza sativa L.)

Heading date of rice is a key agronomic trait determining cultivated areas and seasons and affecting yield. In the present study, ifve primary single segment substitution lines with the same genetic background were used to detect quantitative trait loci （QTLs） for heading date in rice. Two QTLs, qHD3 and qHD6 on the short arm of chromosome 3 and the short arm of chromosome 6, respectively, were identiifed under natural long-day （NLD）. Nineteen secondary single segment substitution lines （SSSLs） and seven double segments pyramiding lines were designed to map the two QTLs and to evaluate their epistatic interaction between them. By overlapping mapping, qHD3 was mapped in a 791-kb interval between SSR markers RM3894 and RM569 and qHD6 in a 1 125-kb interval between RM587 and RM225. Results revealed the existence of epistatic interaction between qHD3 and qHD6 under natural long-day （NLD）. It was also found that qHD3 and qHD6 had signiifcant effects on plant height and yield traits, indicating that both of the QTLs have pleiotropic effects.

The CCT transcriptional activator Ghd2 constantly delays the heading date by upregulating CO3 in rice

CONSTANS,CO-like,and TOC1(CCT)family genes play important roles in regulating heading date,which exerts a large impact on the regional and seasonal adaptation of rice.Previous studies have shown that Grain number,plant height,and heading date2(Ghd2)exhibits a negative response to drought stress by directly upregulating Rubisco activase and exerting a negative effect on heading date.However,the target gene of Ghd2 regulating heading date is still unknown.In this study,CO3 is identified by analyzing Ghd2 ChIP-seq data.Ghd2 activates CO3 expression by binding to the CO3 promoter through its CCT domain.EMSA experiments show that the motif CCACTA in the CO3 promoter was recognized by Ghd2.A comparison of the heading dates among plants with CO3 knocked out or overexpressed and double-mutants with Ghd2 overexpressed and CO3 knocked out shows that CO3 negatively and constantly regulates flowering by repressing the transcription of Ehd1,Hd3a,and RFT1.In addition,the target genes of CO3 are explored via a comprehensive analysis of DAP-seq and RNA-seq data.Taken together,these results suggest that Ghd2 directly binds to the downstream gene CO3,and the Ghd2eCO3 module constantly delays heading date via the Ehd1-mediated pathway.

DHD4,a CONSTANS-like family transcription factor,delays heading date by affecting the formation of the FAC complex in rice

Heading date(or flowering time)is one of the most important agronomic traits in rice,influencing its regional adaptability and crop yield.Many major-effect genes for rice heading date have been identified,but in practice they are difficult to be used for rice molecular breeding because of their dramatic effects on heading date.Genes with minor effects on heading date,which are more desirable for fine-tuning flowering time without significant yield penalty,were seldom reported.In this study,we identified a new minor-effect heading date repressor,Delayed Heading Date 4(DHD4).The dhd4 mutant shows a slightly earlier flowering phenotype without a notable yield penalty compared with wild-type plants under natural long-day conditions.DHD4 encodes a CONSTANS-like transcription factor localized in the nucleus.Molecular,biochemical,and genetic assays show that DHD4 can compete with 14-3-3 to interact with OsFD1,thus affecting the formation of the Hd3a-14-3-3-OsFD1 triprotein FAC complex,resulting in reduced expression of OsMADS14 and OsMADS15,and ultimately delaying flowering.Taken together,these results shed new light on the regulation of flowering time in rice and provide a promising target for fine-tuning flowering time to improve the regional adaptability of rice.

Natural Variation in OsPRR37 Regulates Heading Date and Contributes to Rice Cultivation at a Wide Range of Latitudes

Heading date and photoperiod sensitivity are fundamental traits that determine rice adaptation to a wide range of geographic environments. By quantitative trait locus （QTL） mapping and candidate gene analysis using whole- genome re-sequencing, we found that Oryza sativa Pseudo-Response Regulator37 （OsPRR37; hereafter PRR37） is respon- sible for the Early heading7-2 （EH7-2）/Heading date2 （Hd2） QTL which was identified from a cross of late-heading rice ＇Milyang23 （M23）＇ and early-heading rice ＇H143＇. H143 contains a missense mutation of an invariantly conserved amino acid in the CCT （CONSTANS, CO-like, and TOC1） domain of PRR37 protein. In the world rice collection, different types of nonfunctional PRR37 alleles were found in many European and Asian rice cultivars. Notably, the japonica varieties harboring nonfunctional alleles of both Ghd7/Hd4 and PRR37/Hd2 flower extremely early under natural long-day condi- tions, and are adapted to the northernmost regions of rice cultivation, up to 53~ N latitude. Genetic analysis revealed that the effects of PRR37 and Ghd7 alleles on heading date are additive, and PRR37 down-regulates Hd3a expression to suppress flowering under long-day conditions. Our results demonstrate that natural variations in PRR37/Hd2 and GhdT/ Hd4 have contributed to the expansion of rice cultivation to temperate and cooler regions.

LHD1,an Allele of DTH8/Ghd8,Controls Late Heading Date in Common Wild Rice (Oryza rufipogon)

Flowering at suitable time is very important for plants to adapt to complicated environments and produce their seeds successfully for reproduction. In rice （Oryza rufipogon Griff.） photoperiod regulation is one of the important factors for controlling heading date. Common wild rice, the ancestor of cultivated rice, exhibits a late heading date and a more sensitive photoperiodic response than cultivated rice. Here, through map-based cloning, we identified a major quantitative trait loci （QTL） LHD1 （Late Heading Date 1）, an allele of DTH8/Ghd8, which controls the late heading date of wild rice and encodes a putative HAP3/NF-YB/CBF-A subunit of the CCAAT-box-binding transcription factor. Sequence analysis revealed that several variants in the coding region of LHD1 were correlated with a late heading date, and a further complementary study successfully rescued the phenotype. These results suggest that a functional site for LHD1 could be among those variants present in the coding region. We also found that LHD1 could down-regulate the expression of several floral transition activators such as Ehdl, Hd3a and RFT1 under long-day conditions, but not under short-day conditions. This indicates that LHD1 may delay flowering by repressing the expression of Ehdl, Hd3a and RFT1 under long-day conditions.

Genetic Diversity and Association Analysis for Salinity Tolerance, Heading Date and Plant Height of Barley Germplasm Using Simple Sequence Repeat Markers

The objective of this study was to investigate the genetic diversity of barley accessions. Additionally, association trait analysis was conducted for grain yield under salinity, heading date and plant height. For this purpose, 48 barley genotypes were analyzed with 22 microsatellite simple sequence repeat （SSR） markers. Four of the 22 markers （Bmac316, scssr03907, HVM67 and Bmag770） were able to differentiate all barley genotypes. Cluster and principal coordinate analysis allowed a clear grouping between countries from the same region. The genotypes used in this study have been evaluated for agronomic performance in different environments. Conducting association analysis for grain yield under salinity conditions using TASSEL software revealed a close association of the marker Bmag749 （2H, bin 13） in two different environments with common significant alleles （175, 177）, whereas the HVHOTR1 marker （2H, bin 3） was only significant in Sakhar Egypt with alleles size being 158 and 161. Heading date also showed an association with scssr03907 through the common significant specific allele 111 and EBmac0415 markers in three different agro climatic locations, whereas HVCMA, scssr00103 and HVM67 were linked to heading date in the Egyptian environment only. The plant height association analysis revealed significant markers Bmag770 via the significant allele 152 and scssr09398.

Down-Regulation of OsGRF1 Gene in Rice rhdl Mutant Results in Reduced Heading Date

Abstract: A rice mutant with reduced heading date (designated rhd1) found in a transgenic line of cultivar Teqing 2 (Oryza sativa L. ssp. indica) was used to identify the genes related to rice heading and thereby to study its molecular mechanism. Genetic analysis showed that rhd1 was a dominant mutation and did not result from T-DNA insertion. By using the differential display polymerase chain reaction (DD-PCR) technique, differential gene expression between rhd1 and Teqing 2 was compared and a rhd1-down-regulated c DNA fragment was identified. Sequence analysis showed that this fragment shared 99% similarity to the OsGRF1 (O. sativa growth-regulating factor 1) gene. The OsGRF1 gene encodes a putative transcription factor, which contains two conserved regions: the QLQ (Gln, Leu, Gln) and WRC (Trp, Arg, Cys) domains. Southern analysis indicates that OsGRF1 may be encoded by single copy gene in the rice genome. RNA interference results revealed that transgenic lines with reduced OsGRF1 transcript displayed delayed growth and development, developed small leaves, and had delayed heading. The extent of the phenotypes developed was well-correlated with the OsGRF1 gene transcript. Our results clearly demonstrate that the OsGRF1 gene is not only involved in regulating growth at the juvenile stage, but that it may also be involved in the regulation of heading in rice.

The Distribution of Japonica Rice Cultivars in the Lower Region of the Yangtze River Valley is Determined by Its Photoperiod-sensitivity and Heading Date Genotypes

There are generally four recognized classes of japonica rice cultivars grown in the lower region of the Yangtze River valley. The geographical distribution of the four classes is latitude-dependent. Variation for heading date （HD） among 29 japonica rice cultivars grown in the lower region of the Yangtze River valley and belonging to the four classes was characterized, and their sensitivity to variations in photoperiod and temperature was analyzed. All of the cultivars were sensitive to both photoperiod and temperature. A regression analysis showed that HD is closely correlated with photoperiod sensitivity （PS）. The PS of the four classes increased gradually from the medium maturing middle （MMM） types, through the late maturing middle （LMM） and early maturing late （EML） types to the medium maturing late （MML） types. Crosses with tester lines established that almost all of the cultivars carry the dominant early-heading allele at Ef-1, the photoperiod insensitive allele e2 and the PS alleles E1 or E1^t. Most of the MMM, LMM and MML types carry the insensitive allele e3, while EML types have either E3 or E3^t. At Se-1, MMM and LMM types have Se-1^e, some EML types have Se-1^e and others Se-1^n, while the MML types are mostly Se-fn. The PS of some MMM, LMM and EML types is reduced by the presence of hd2. These results show that the distribution of the four rice cultivar classes from high latitude to low latitude regions depended on a gradual increase in PS, which is mainly determined by its HD genotypes.

Genotypes of Heading Date of Middle Indica Rice in the Mid-lower Region of the Yangtze River

Middle indica cultivars are planted in the middle to lower regions of the Yangtze River. Hybrid combinations with these cultivars have a high yield potential but the presence of late-transgressive-segregants limits the further exploitation of heterosis. To understand the genetic basis of the heading date in these middle maturing cultivars, we carried out a genetic analysis of 10 typical middle-season cultivars using a number of heading date isogenic lines under both long and short day conditions. The results showed that Teqing, 752, CDR22, Bo B, 9311 and 11-32B carry two photoperiod sensitive genes E1 or Se.1 and E3, and Teqing, 752 and CDR22 carry a dominant early-heading gene Ef-1 while Bo B, 9311 and 11-32B carry a recessive late-heading gene ef-1. Based on the findings of the present and previous works on Guichao 2, Minghui 63, Nanjing 11 and Pei＇ai64S, it was concluded that all of these middle indica cultivars carried the recessive allele hd2, which could inhibit the expression of El or Se.1, and they formed rational combinations of genotype for heading date during a long period of evolution. The effects of the different combinations of genotype for heading date on rice cultivation and extension were discussed, and genetic basis of broad adaptability of hybrid middle indica cultivars was analyzed.

Hd1,Ghd7,and DTH8 synergistically determine the rice heading date and yield-related agronomic traits

Heading date determines the seasonal and regional adaptation of rice(Oryza sativa L.)varieties and is mainly controlled by photoperiod sensitivity(PS).The core heading date genes Hd1,Ghd7,DTH8,and PRR37 act synergistically in regulating the PS.In this study,we systematically analyze the heading date,PS,and agronomic traits of eight homozygous lines with various combinations of Hd1,Ghd7,and DTH8 alleles in the prr37 background under long-day(LD)and short-day(SD)conditions,respectively.We find that Hd1 alone promotes heading,regardless of the day length.However,under LDs,Hd1 suppresses flowering,in coordination with functional Ghd7 or with Ghd7 and DTH8.These loci cooperate to negatively regulate the Ehd1-Hd3 a/RFT1 pathway and delay heading.Under SDs,Hd1 competes with various heading suppressors to promote heading.Therefore,the dual function of Hd1 is vital for PS.The lines carrying Hd1 alone show reduced plant height with fewer primary and secondary branches in panicles.Lines carrying Ghd7 and DTH8(with hd1)show delayed heading and improve agronomic traits.Overall,our results reveal the regulation of rice PS flowering by the core heading date genes and their effects on agronomic traits,providing valuable information for the selection of rice varieties for adaptation to different light and temperature conditions.

Two Novel QTLs for Heading Date Are Identified Using a Set of Chromosome Segment Substitution Lines in Rice(Oryza sativa L.)

Heading date in rice is a typical quantitative trait controlled by multiple quantitative trait loci （QTLs）. It is mainly regulated by environmental factors such as photoperiod and temperature （Izawa, 2007）. Many QTLs for heading date have been identified using different mapping populations and methods （http：// www.gramene.org/qtl）. Up to date, several major heading date QTLs have been cloned by map-based cloning strategy （Yano et al., 2000; Takahashi et al., 2001; Kojima et al., 2002; Doi et al., 2004; Xue et al., 2008; Wei et al., 2010; Yan et al.,