A 48-bp deletion upstream of LIGULELESS 1 alters rice panicle architecture

Panicle architecture is an agronomic determinant of crop yield and a target for cereal crop improvement.To investigate its molecular mechanisms in rice,we performed map-based cloning and characterization of OPEN PANICLE 1(OP1),a gain-of-function allele of LIGULELESS 1(LG1),controlling the spread-panicle phenotype.This allele results from a 48-bp deletion in the LG1 upstream region and promotes pulvinus development at the base of the primary branch.Increased OP1 expression and altered panicle phenotype in chimeric transgenic plants and upstream-region knockout mutants indicated that the deletion regulates spread-panicle architecture in the mutant spread panicle 1(sp1).Knocking out BRASSINOSTEROID UPREGULATED1(BU1)gene in the background of OP1 complementary plants resulted in compact panicles,suggesting OP1 may regulate inflorescence architecture via the brassinosteroid signaling pathway.We regard that manipulating the upstream regulatory region of OP1 or genes involved in BR signal pathway could be an efficient way to improve rice inflorescence architecture.

Haploids can be induced in knockout mutants of OsPLA1,but not OsDMP3 or OsDMP6,in rice

Doubled haploid(DH)technology is an important tool in crop breeding because it can significantly accelerate the breeding process.ZmPLA1/MATL/NLD and ZmDMP are two key genes controlling haploid induction(HI)in maize,exhibiting a synergistic effect.However,it is unknown whether knock out of ZmDMP orthologs can stimulate HI in rice.In this study,a ZmPLA1 ortholog(OsPLA1)and two ZmDMP orthologs(OsDMP3 and OsDMP6)were identified in rice.All three genes encode plasma membrane-localized proteins and were highly expressed in mature anthers.Knockout of OsPLA1 in both Minghui 63 and Nipponbare resulted in reduced seed setting rate(SSR)and caused HI.The osdmp3,osdmp6 and the double mutant failed to trigger HI independently,nor increased the haploid induction rate(HIR)when combined with ospla1.Repeated pollinations operations of QX654A with the ospla1 mutant significantly improve SSR,while reducing HIR.RNA-seq profiling of mature ospla1 mutant anthers indicated that a large number of differentially expressed genes(DEGs)were enriched in redox homeostasis and lipid metabolic GO terms,plant hormone signal transduction,and MAPK signaling pathways.These findings provide important insights towards construction of an efficient DH breeding technology and study of the molecular mechanism of HI in rice.

A gain-of-function mutation of OsMAPK6 leads to long grain in rice

Grain size is one of the most important agronomic traits controlling grain yield. Development of novel germplasm with large grains would be beneficial for crop improvement. We report the genetic identification and functional analysis of the LONG GRAIN 6(LOG6) gene, which is identical to MITOGENACTIVATED PROTEIN KINASE 6(OsMAPK6), affecting grain length of rice. Map-based cloning revealed that the long-grain phenotype of log6-D results from a glutamine(E) to lysine(K) mutation in the conserved TEY motif of OsMAPK6. In near-isogenic lines(NILs), the log6-D allele increased grain length and grain yield of Guichao 2(GC2), Teqing(TQ), and 93–11. Sequence analysis revealed 10 OsMAPK6 haplotypes,with xian(indica) and geng(japonica) harboring different haplotypes. Our findings shed light on the function of MAPKs and offer a novel dominant allele for improving the grain yield of rice.

Identification and mapping of genes for improving yield from Chinese common wild rice (O.rufipogon Griff.) using advanced backcross QTL analysis

To identify useful genes from wild rice which have been lost or weakened in cultivated rice has become more and more important for modern breeding strategy. In this study, a BC4 population derived from 94W1, an accession of common wild rice (Oryza rufipogon Griff.) from Dongxiang in Jiangxi Province of China, as the donor, and a high-yielding Indica cultivar (O. sativa L.), 'Guichao 2', as the recipient, was used to identify quantitative trait loci (QTL) associated with yield and its components. Based on the analysis for the genotype of BC4F1 population with 87 SSR markers distributed throughout the genome and investigation of the plant height, yield and yield components of BC4F2, a total of 52 QTLs, were detected. Of 7 QTLs associated with grain yield per plant, 2 QTLs on chromosome 2 and chromosome 11 for grain yield, explaining 16% and 11% of the phenotypic variance respectively, were identified. The alleles from Dongxiang common wild rice in those two loci could increase the yield of 'Guichao 2' by

GS6, A Member of the GRAS Gene Family,Negatively Regulates Grain Size in Rice

Grain size is an important yield-related trait in rice. Intensive artificial selection for grain size during domestication is evidenced by the larger grains of most of today＇s cultivars compared with their wild relatives. However, the molecular genetic control of rice grain size is still not well characterized. Here, we report the identification and cloning of Grain Size 6 （GS6）, which plays an important role in reducing grain size in rice. A premature stop at the ＋348 position in the coding sequence （CDS） of GS6 increased grain width and weight significantly. Alignment of the CDS regions of GS6 in 90 rice materials revealed three GS6 alleles. Most japonica varieties （95%） harbor the Type I haplotype, and 62.9% of indica varieties harbor the Type II haplotype. Association analysis revealed that the Type I haplotype tends to increase the width and weight of grains more than either of the Type II or Type III haplotypes. Further investigation of genetic diversity and the evolutionary mechanisms of GS6 showed that the GS6 gene was strongly selected in japonica cultivars. In addition, a ＂ggc＂ repeat region identified in the region that encodes the GRAS domain of GS6 played an important historic role in the domestication of grain size in rice. Knowledge of the function of GS6 might aid efforts to elucidate the molecular mechanisms that control grain development and evolution in rice plants, and could facilitate the genetic improvement of rice yield.

NARROW AND ROLLED LEAF 2 regulates leaf shape,male fertility, and seed size in rice

Grain yield in rice （Oryza sativa L.） is closely related to leaf and flower development. Coordinative regulation of leaf, pollen, and seed development in rice as a critical biological and agricultural question should be addressed. Here we identified two allelic rice mutants with narrow and semi- rolled leaves, named narrow and rolled leaf2-1 （nrl2-1） and nr12- 2. Map-based molecular cloning revealed that NRL2 encodes a novel protein with unknown biochemical function. The mutation of NRL2 caused pleiotropic effects, including a reduction in the number of longitudinal veins, defective abaxial sclerenchymatous cell differentiation, abnormal tape- tum degeneration and microspore development, and the formation of more slender seeds compared with the wild type （WT）. The NRL2 protein interacted with Rolling-leaf （RL14）, causing the leaves of the nrl2 mutants to have a higher cellulose content and lower Iignin content than the WT, which may have been related to sclerenchymatous cell differentia- tion and tapetum degeneration. Thus, this gene is an essential developmental regulator controlling fundamental cellular and developmental processes, serving as a potential breeding target for high-yielding rice cultivars.

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.

Additive and Over-dominant Effects Resulting from Epistatic Loci Are the Primary Genetic Basis of Heterosis in Rice

A set of 148 F9 recombinant inbred lines （RILs） was developed from the cross of an indica cultivar 93-11 and japonica cultivar DTI13, showing strong F1 heterosis. Subsequently, two backcross F1 （BCFI） populations were constructed by backcrossing these 148 RILs to two parents, 93-11 and DT713. These three related populations （281BCF1 lines, 148 RILs） were phenotyped for six yield-related traits in two locations. Significant inbreeding depression was detected in the population of RILS and a high level of heterosis was observed in the two BCF1 populations. A total of 42 main-effect quantitative trait loci （M-QTLs） and 109 epistatic effect QTL pairs （E-QTLs） were detected in the three related populations using the mixed model approach. By comparing the genetic effects of these QTLs detected in the RILs, BCF1 performance and mid-parental heterosis （HMp）, we found that, in both BCF1 populations, the QTLs detected could be classified into two predominant types： additive and over-dominant loci, which indicated that the additive and over-dominant effect were more important than complete or partially dominance for M-QTLs and E-QTLs. Further, we found that the E-QTLs detected collectively explained a larger portion of the total phenotypic variation than the M-QTLs in both RILs and BCF1 populations. All of these results suggest that additive and over-dominance resulting from epistatic loci might be the primary genetic basis of heterosis in rice.

Natural Variations at TIG1 Encoding a TCP Tran scription Factor Contribute to Plant Architecture Domestication in Rice

The modification of plant architecture is a crucial target in rice domestication and modern genetic improvement.Although several genes regulating rice plant architecture have been characterized,the molecular mechanisms underlying rice plant architecture domestication remain largely unclear.Here we show that the inclined tiller growth in wild rice is controlled by a single dominant gene,TILLER INCLINED GROWTH 1 (T/Gf),which is located on chromosome 8 and encodes a TCP transcriptional activator.TIG1 is primarily expressed in the adaxial side of the tiller base,promotes cell elongation,and enlarges the tiller angle in wild rice.Variations in the TIG1 promoter of indica cultivars {tig1 allele) resulted in decreased expression of TIG1 in the adaxial side of tiller base and reduced cell length and tiller angle,leading to the transition from inclined tiller growth in wild rice to erect tiller growth during rice domestication.TIG1 positively regulates the expression of EXP A3,EXPB5,and SAUR39 to promote cell elongation and increase the tiller angle.Selective sweep analysis revealed that the tig1 allele was selected in indica cultivars by human beings.The cloning and characterization of TIG1 supports a new scenario of plant architecture evolution in rice.

Microarray-Assisted Fine-Mapping of Quantitative Trait Loci for Cold Tolerance in Rice

Many important agronomic traits, including cold stress resistance, are complex and controlled by quantitative trait loci （QTLs）. Isolation of these QTLs will greatly benefit the agricultural industry but it is a challenging task. This study explored an integrated strategy by combining microarray with QTL-mapping in order to identify cold-tolerant QTLs from a cold-tolerant variety ILl12 at early-seedling stage. All the early seedlings of IL112 survived normally for 9 d at 4-5℃, while Guichao2 （GC2）, an indica cultivar, died after 4 d under the same conditions. Using the F2-3 population derived from the progeny of GC2 and ILl12, we identified seven QTLs for cold tolerance. Furthermore, we performed Affymetrix rice whole-genome array hybridization and obtained the expression profiles of ILl12 and GC2 under both low-temperature and normal conditions. Four genes were selected as cold QTL-related candidates, based on microarray data mining and QTL-mapping. One candidate gene, LOC_Os07g22494, was shown to be highly associated with cold tolerance in a number of rice varieties and in the F2-3 population, and its overexpression transgenic rice plants displayed strong tolerance to low temperature at early-seedling stage. The results indicated that overexpression of this gene （LOC_Os07g22494） could increase cold tolerance in rice seedlings. Therefore, this study provides a promising strategy for identifying candidate genes in defined QTL regions.

Molecular Evolution of the TACl Gene from Rice(Oryza sativa L.)

Tiller angle is a key feature of the architecture of cultivated rice （Oryza sativa）, since it determines planting density and influences rice yield. Our previous work identified Tiller Angle Control 1 （TACI） as a major quantitative trait locus that controls rice filler angle. To further clarify the evolutionary characterization of the TAC1 gene, we compared a TACl-containing 3164-bp genomic region among 113 cultivated varieties and 48 accessions of wild rice, including 43 accessions of O. rufipogon and five accessions of O. nivara. Only one single nucleotide polymorphism （SNP）, a synonymous substitution, was detected in TAC1 coding regions of the cultivated rice varieties, whereas one synonymous and one nonsynonymous SNP were detected among the TAC1 coding regions of wild rice accessions. These data indicate that little natural mutation and modification in the TAC1 coding region occurred within the cultivated rice and its progenitor during evolution. Nucleotide diversities in the TAC1 gene regions of O. sativa and O. rufipogon of 0.00116 and 0.00112, respectively, further indicate that TAC1 has been highly conserved during the course of rice domestication. A functional nucleotide polymorphism （FNP） of TAC1 was only found in the japonica rice group. A neutrality test revealed strong selection, especially in the 3＇-flanking region of the TAC1 coding region containing the FNP in the japonica rice group. However, no selection occurred in the indica and wild-rice groups. A phylogenetic tree derived from TAC1 sequence analysis suggests that the indica and japonica subspecies arose indepen- dently during the domestication of wild rice.

Analysis of QTLs for yield-related traits in Yuanjiang common wild rice (Oryza rufipogon Griff.)

Using an accession of common wild rice （Oryza rufipogon Griff.） collected from Yuanjiang County, Yunnan Province, China, as the donor and an elite cultivar 93-11, widely used in two-line indica hybrid rice production in China, as the recurrent parent, an advanced backcross populations were developed. Through genotyping of 187 SSR markers and investigation of six yield-related traits of two gen- erations （BC4F2 and BC4F4）, a total of 26 QTLs were detected by employing single point analysis and interval mapping in both genera-tions. Of the 26 QTLs, the alleles of 10 （38.5%） QTLs originating from O. rufipogon had shown a beneficial effect for yield-related traits in the 93-11 genetic background. In addition, five QTLs controlling yield and its components were newly identified, indicating that there are potentially novel alleles in Yuanjiang common wild rice. Three regions underling significant QTLs for several yield-related traits were detected on chromosome 1, 7 and 12. The QTL clusters were founded and corresponding agronomic traits of those QTLs showed highly significant correlation, suggesting the pleiotropism or tight linkage. Fine-mapping and cloning of these yield-related QTLs from wild rice would be helpful to elucidating molecular mechanism of rice domestication and rice breeding in the future.

Polyamine oxidase 3 is involved in salt tolerance at the germination stage in rice

Soil salinity inhibits seed germination and reduces seedling survival rate,resulting in significant yield reductions in crops.Here,we report the identification of a polyamine oxidase,OsPAO3,conferring salt tolerance at the germination stage in rice(Oryza sativa L.),through map-based cloning approach.OsPAO3 is up-regulated under salt stress at the germination stage and highly expressed in various organs.Overexpression of OsPAO3 increases activity of polyamine oxidases,enhancing the polyamine content in seed coleoptiles.Increased polyamine may lead to the enhance of the activity of ROS-scavenging enzymes to eliminate over-accumulated H;O;and to reduce Na;content in seed coleoptiles to maintain ion homeostasis and weaken Na;damage.These changes resulted in stronger salt tolerance at the germination stage in rice.Our findings not only provide a unique gene for breeding new salt-tolerant rice cultivars but also help to elucidate the mechanism of salt tolerance in rice.

The genetic control of glabrous glume during African rice domestication

African cultivated rice,Oryza glaberrima,is characterized by its glabrous glumes.During domestication,the pubescent glumes of its wild ancestor,Oryza barthii,lost their trichomes,and in this study,we show that glabrous glume 5(GLAG5),a WUSCHEL-like homeobox transcription factor gene on chromosome 5,is required for trichome development.DNA methylation associated with an hATtransposable element inserted in the promoter region of GLAG5 is found to reduce its expression,leading to the formation of glabrous glumes and leaves in African cultivated rice.Among 82 African cultivated rice varieties investigated in this study,59(approximately 71%)lines exhibit glabrous glumes and harbor the hAT transposon;however,the other 23 varieties(approximately 29%),which exhibit pubescent glumes,lack the hAT transposon,indicating that glag5 had undergone strong artificial selection.Theπ;/π;ratios also show the hAT transposon insertions influence the genetic diversity of an approximately 150-kb interval encompassing the GLAG5 locus.The identification of the GLAG5 gene provides new insights into the domestication of cultivated rice in Africa.We speculate that the selection of varieties with mutations in their promoter regions is an important aspect of crop domestication.