Hawthorn leaf flavonoids alleviate the deterioration of atherosclerosis by inhibiting SCAP-SREBP2-LDLR pathway through sPLA2-â ¡A signaling in macrophages in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Hawthorn leaves are a combination of the dried leaves of the Rosaceae plants, i.e., Crataegus pinnatifida Bge. or Crataegus pinnatifida Bge. var. major N. E. Br., is primarily cultivated in East Asia, North America, and Europe. hawthorn leaf flavonoids (HLF) are the main part of extraction. The HLF have demonstrated potential in preventing hypertension, inflammation, hyperlipidemia, and atherosclerosis. However, the potential pharmacological mechanism behind its anti-atherosclerotic effect has yet to be explored. AIM OF THE STUDY: The in vivo and in vitro effects of HLF on lipid-mediated foam cell formation were investigated, with a specific focus on the levels of secreted phospholipase A2 type IIA (sPLA2-II A) in macrophage cells. MATERIALS AND METHODS: The primary constituents of HLF were analyzed using ultra-high performance liquid chromatography and liquid chromatography-tandem mass spectrometry. In vivo, HLF, at concentrations of 5 mg/kg, 20 mg/kg, and 40 mg/kg, were administered to apolipoprotein E knockout mice (ApoE-/-) fed by high-fat diet (HFD) for 16 weeks. Aorta and serum samples were collected to identify lesion areas and lipids through mass spectrometry analysis to dissect the pathological process. RAW264.7 cells were incubated with oxidized low-density lipoprotein (ox-LDL) alone, or ox-LDL combined with different doses of HLF (100, 50, and 25 µg/ml), or ox-LDL plus 24-h sPLA2-IIA inhibitors, for cell biology analysis. Lipids and inflammatory cytokines were detected using biochemical analyzers and ELISA, while plaque size and collagen content of plaque were assessed by HE and the Masson staining of the aorta. The lipid deposition in macrophages was observed by Oil Red O staining. The expression of sPLA2-IIA and SCAP-SREBP2-LDLR was determined by RT-qPCR and Western blot analysis. RESULTS: The chemical profile of HLF was studied using UPLC-Q-TOF-MS/MS, allowing the tentative identification of 20 compounds, comprising 1 phenolic acid, 9 flavonols and 10 flavones, including isovitexin, vitexin-4″-O-glucoside, quercetin-3-O-robibioside, rutin, vitexin-2″-O-rhamnoside, quercetin, etc. HLF decreased total cholesterol (TC), triglycerides (TG), low-density lipoprotein cholesterol (LDL-C), and non-high-density lipoprotein cholesterol (non-HDL-C) levels in ApoE-/- mice (P < 0.05), reduced ox-LDL uptake, inhibited level of inflammatory factors, such as IL-6, IL-8, TNF-α, and IL-1ꞵ (P < 0.001), and alleviated aortic plaques with a thicker fibrous cap. HLF effectively attenuated foam cell formation in ox-LDL-treated RAW264.7 macrophages, and reduced levels of intracellular TC, free cholesterol (FC), cholesteryl ester (CE), IL-6, TNF-α, and IL-1ß (P < 0.001). In both in vivo and in vitro experiments, HLF significantly downregulated the expression of sPLA2-IIA, SCAP, SREBP2, LDLR, HMGCR, and LOX-1 (P < 0.05). Furthermore, sPLA2-IIA inhibitor effectively mitigated inflammatory release in RAW264.7 macrophages and regulated SCAP-SREBP2-LDLR signaling pathway by inhibiting sPLA2-IIA secretion (P < 0.05). CONCLUSION: HLF exerted a protective effect against atherosclerosis through inhibiting sPLA2-IIA to diminish SCAP-SREBP2-LDLR signaling pathway, to reduce LDL uptake caused foam cell formation, and to slow down the progression of atherosclerosis in mice.

Sex differences in antioxidant ability and energy metabolism level resulting in the difference of hypoxia tolerance in red swamp crayfish (Procambarus clarkii).

Sexual dimorphism widely exists in crustaceans. However, sex differences in the hypoxia tolerance of crayfish have rarely been reported. In this study, the differences in hypoxia tolerance between the two sexes of crayfish were assessed according to mortality, pathological features of hepatopancreas, antioxidant enzyme activity and differentially expressed genes (DEGs) analysis using transcriptome. The results showed that male crayfish displayed significantly higher mortality than the female under hypoxia stress (p < 0.05). Furthermore, female crayfish demonstrated higher levels of antioxidant enzyme activity. Hematoxylin-eosin staining analysis revealed that the damage of hepatopancreas was more severe in the male crayfish compared to the female crayfish. Additionally, there was higher expression level of the DEGs in hypoxia-inducible factor (HIF) pathway and higher energy metabolism level in the female compared to the male. Together, these findings suggest that the female crayfish with higher antioxidant ability and energy metabolism level exhibits stronger hypoxia tolerance than the male crayfish, providing the theoretical support for investigating sex differences in hypoxia tolerance among crustaceans.

Trained immunity can improve the disease resistance of red swamp crayfish (Procambarus clarkii).

Trained immunity has been widely observed in mammals. Its identification in red swamp crayfish (Procambarus clarkii) is important for disease resistance in the crayfish farming industry. In this study, the mortality, expression of immune genes, production of reactive oxygen species (ROS), and phagocytosis ability of haemocytes in crayfish infected by pathogens (Vibrio parahaemolyticus or white spot syndrome virus) and crayfish trained with ß-glucan or PBS (the control) were assessed when they were re-challenged by the pathogens. The results showed that the mortality of the trained and re-challenged crayfish were significantly lower than those of the untrained and challenged crayfish. Furthermore, the expression of immune genes, including Resistance (R), ALF, crustin2, and proPO, ROS levels, and phagocytosis ability of haemocytes, was significantly improved in the trained crayfish compared to that in the untrained crayfish. Interestingly, we found that the immune memory of trained crayfish lasted for at least 18 days. Together, these results indicate that crayfish develops trained immunity that can play an important role in the disease resistance. This suggests that trained immunity may be applied to improve disease resistance and crayfish production.

Robust strategy for disease resistance and increasing production breeding in red swamp crayfish (Procambarus clarkii).

Red swamp crayfish (Procambarus clarkii) is an important aquaculture species in China. With increasing crayfish culture, a number of outbreaks of various diseases have been identified in crayfish. Despite this, there are no reports on the application of disease resistance genes in the molecular breeding of crayfish. In this study, transcriptome analysis was performed to explore the disease resistance genes in crayfish, with a focus on investigating the genetic variations in the open reading frames of these genes, for subsequent haplotype analysis. Furthermore, pathogen-challenge experiments were carried out in the crayfish, to identify the favoured haplotypes. A novel disease resistance gene, R (Resistance), was identified by means of transcriptome analysis. In total, two, four, and five haplotypes of the three disease resistance genes, ALF, R, and crustin2, respectively, were detected. ALF1, R1, and Cru1 were the favoured haplotypes of ALF, R, and crustin2, respectively. Subsequently, the favoured haplotype combinations of the different genes were obtained, and a series of molecular markers were developed to identify them. Finally, we propose a molecular breeding strategy to enhance the disease resistance of crayfish, and thus, improve its production.

A minor QTL, SG3, encoding an R2R3-MYB protein, negatively controls grain length in rice.

KEY MESSAGE: SG3, an R2R3 MYB protein coding gene that tightly linked to a major QTLGS3, negatively regulates grain length while dependent on the status ofGS3in rice. It is often very difficult to isolate a minor QTL that is closely linked to a major QTL in rice. In this study, we focused on the isolation of a minor grain length QTL, small grain 3 (SG3), which is closely linked to the major QTL grain size 3 (GS3). The genetic effect of SG3 on grain length was dependent on GS3 status. Its genetic effect was larger in the presence of nonfunctional sg3 than functional SG3. A large number of near-isogenic F2 plants in which GS3 was fixed with nonfunctional alleles were developed to clone SG3. A gene encoding an R2R3 MYB domain transcriptional regulator was identified as the candidate gene for SG3. SG3 overexpression and knockdown plants showed shortened and elongated grains, respectively, which demonstrated that SG3 acts as a negative regulator of grain length. SG3 was preferentially expressed in panicles after flowering, and SG3 acted as a transcription activator. Comparative sequencing analysis identified a 12-bp insertion in the third exon of NYZ that led to a frameshift and resulted in a premature stop codon. The insertion/deletion was associated with grain length in the presence of functional GS3 in the indica subspecies. SG3 and GS3 were frequently in coupling phase in indica rice, making them good targets for the breeding of cultivars with short or long grains. The isolation of the SG3 MYB gene provides new gene resource and contributes to the regulatory network of grain length in rice.

Wide Grain 7 increases grain width by enhancing H3K4me3 enrichment in the OsMADS1 promoter in rice (Oryza sativa L.).

Grain size is a major determinant of grain weight, a key component of grain yield of rice. Here, we identified the grain size gene WIDE GRAIN 7 (WG7) from a T-DNA insertion mutant. The grain size of WG7 knockout mutants and WG7 overexpression lines indicated that WG7 is a positive regulator of grain size. WG7 encodes a cysteine-tryptophan (CW) domain-containing transcriptional activator. EMSAs and ChIP-qPCR assay confirmed that WG7 directly bound to the promoter of OsMADS1, a grain size gene, and thereby significantly activated its expression. Point mutations showed that the cis-element CATTTC motif in the promoter was the binding site of WG7. Compared with the wild-type, deletion mutants of the cis-element motif exhibited lower expression of OsMADS1 and produced narrower grains, implicating the requirement of this motif for WG7 function. ChIP-qPCR assays showed that WG7 enhanced histone H3K4me3 enrichment in the promoter of OsMADS1. WG7 underwent directional selection due to the poor fertility of the non-functional mutant. These findings demonstrated that WG7 upregulated OsMADS1 expression by directly binding to its promoter, enhanced histone H3K4me3 enrichment in the promoter and ultimately increased grain width. This study will enrich the knowledge concerning the regulatory network of grain size formation in rice.

Genome-wide dissection of segregation distortion using multiple inter-subspecific crosses in rice.

Mendelian inheritance can ensure equal segregation of alleles from parents to offspring, which provides fundamental basis for genetics and molecular biology. Segregation distortion (SD) leads to preferential transmission of certain alleles from generation to generation. Such violation of Mendelian genetic principle is often accompanied by reproductive isolation and eventually speciation. Although SD is observed in a wide range of species from plants to animals, genome-wide dissection of such biased transmission of gametes is rare. Using nine inter-subspecific rice crosses, a genome-wide screen for SD loci is performed, which reveals 61 single-locus quantitative trait loci and 194 digenic interactions showing distorted transmission ratio, among which 24 new SD loci are identified. Biased transmission of alleles is observed in all nine crosses, suggesting that SD exists extensively in rice populations. 72.13% distorted regions are repeatedly detected in multiple populations, and the most prevalent SD hotspot that observed in eight populations is mapped to chromosome 3. Xian alleles are transmitted at higher frequencies than geng alleles in inter-subspecific crosses, which change the genetic composition of the rice populations. Epistatic interaction contributes significantly to the deviation of Mendelian segregation at the whole-genome level in rice, which is distinct from that in animals. These results provide an extensive archive for investigating the genetic basis of SD in rice, which have significant implications in understanding the reproductive isolation and formation of inter-subspecific barriers during the evolution.

Short Panicle 3 controls panicle architecture by upregulating APO2/RFL and increasing cytokinin content in rice.

Inflorescence architecture is a major determinant of spikelet numbers per panicle, a key component of grain yield in rice. In this study, Short Panicle 3 (SP3) was identified from a short panicle 3 (sp3) mutant in which T-DNA was inserted in the promoter of SP3, resulting in a knockdown mutation. SP3 encodes a DNA binding with one finger (Dof) transcriptional activator. Quantitative real time (qRT)-PCR and RNA in situ hybridization assays confirmed that SP3 is preferentially expressed in the young rice inflorescence, specifically in the branch primordial regions. SP3 acts as a negative regulator of inflorescence meristem abortion by upregulating APO2/RFL. SP3 both up- and downregulates expression of genes involved in cytokinin biosynthesis and catabolism, respectively. Consequently, cytokinin concentrations are decreased in young sp3 panicles, thereby leading to small panicles having fewer branches and spikelets. Our findings support a model in which SP3 regulates panicle architecture by modulating cytokinin homeostasis. Potential applications to rice breeding, through gene-editing of the SP3 promoter are assessed.

Minor Effects of 11 Dof Family Genes Contribute to the Missing Heritability of Heading Date in Rice (Oryza sativa L.).

DNA binding with one finger (Dof) proteins are plant-specific transcription factors with important and diverse functions in seed germination, flowering time, and biotic and abiotic stresses. In this study, haplotype-based association analysis was conducted between heading date and 30 Dof family genes in a worldwide germplasm collection. Of these, 22 Dof genes were associated with heading date. Multiple comparisons among haplotypes revealed their diverse functions in promoting and suppressing heading date under short-day (SD) and long-day (LD) conditions. They cumulatively made a considerable contribution to the missing heritability of heading date. A set of knockout mutants of 30 Dof genes generated by CRISPR/Cas9-mediated genome editing technology showed that 11 and 9 Dof genes regulated heading date under LD and SD, respectively. Phenotype measurement of mutants showed that these 11 and 9 Dof genes slightly regulated heading with effects of 2-5 days under LD and SD, respectively. Both mutant and natural variation assays indicated functional redundancy in regulating heading date among Dof family genes. Nucleotide diversity analysis suggested that most Dof genes have been subjected to selection during domestication and improvement. Beyond heading date, this set of mutants is also a good resource for evaluating the function of Dof genes in regulating stress tolerance and seed germination.

OsMFT1 increases spikelets per panicle and delays heading date in rice by suppressing Ehd1, FZP and SEPALLATA-like genes.

Heading date and panicle architecture are important agronomic traits in rice. Here, we identified a gene MOTHER OF FT AND TFL1 (OsMFT1) that regulates rice heading and panicle architecture. Overexpressing OsMFT1 delayed heading date by over 7 d and greatly increased spikelets per panicle and the number of branches. In contrast, OsMFT1 knockout mutants had an advanced heading date and reduced spikelets per panicle. Overexpression of OsMFT1 significantly suppressed Ehd1 expression, and Ghd7 up-regulated OsMFT1 expression. Double mutants showed that OsMFT1 acted downstream of Ghd7. In addition, transcription factor OsLFL1 was verified to directly bind to the promoter of OsMFT1 via an RY motif and activate the expression of OsMFT1 in vivo and in vitro. RNA-seq and RNA in situ hybridization analysis confirmed that OsMFT1 repressed expression of FZP and five SEPALLATA-like genes, indicating that the transition from branch meristem to spikelet meristem was delayed and thus more panicle branches were produced. Therefore, OsMFT1 is a suppressor of flowering acting downstream of Ghd7 and upstream of Ehd1, and a positive regulator of panicle architecture.

A Rice Genetic Improvement Boom by Next Generation Sequencing.

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

Duplication of an upstream silencer of FZP increases grain yield in rice.

Transcriptional silencer and copy number variants (CNVs) are associated with gene expression. However, their roles in generating phenotypes have not been well studied. Here we identified a rice quantitative trait locus, SGDP7 (Small Grain and Dense Panicle 7). SGDP7 is identical to FZP (FRIZZY PANICLE), which represses the formation of axillary meristems. The causal mutation of SGDP7 is an 18-bp fragment, named CNV-18bp, which was inserted ~5.3 kb upstream of FZP and resulted in a tandem duplication in the cultivar Chuan 7. The CNV-18bp duplication repressed FZP expression, prolonged the panicle branching period and increased grain yield by more than 15% through substantially increasing the number of spikelets per panicle (SPP) and slightly decreasing the 1,000-grain weight (TGW). The transcription repressor OsBZR1 binds the CGTG motifs in CNV-18bp and thereby represses FZP expression, indicating that CNV-18bp is the upstream silencer of FZP. These findings showed that the silencer CNVs coordinate a trade-off between SPP and TGW by fine-tuning FZP expression, and balancing the trade-off could enhance yield potential.

Genome-Wide Association Analysis Reveals Different Genetic Control in Panicle Architecture Between and Rice.

Panicle architecture determines the number of spikelets per panicle (SPP) and is highly associated with grain yield in rice ( L.). Understanding the genetic basis of panicle architecture is important for improving the yield of rice grain. In this study, we dissected panicle architecture traits into eight components, which were phenotyped from a germplasm collection of 529 cultivars. Multiple regression analysis revealed that the number of secondary branch (NSB) was the major factor that contributed to SPP. Genome-wide association analysis was performed independently for the eight particle architecture traits observed in the and rice subpopulations compared with the whole rice population. In total, 30 loci were associated with these traits. Of these, 13 loci were closely linked to known panicle architecture genes, and 17 novel loci were repeatedly identified in different environments. An association signal cluster was identified for NSB and number of spikelets per secondary branch (NSSB) in the region of 31.6 to 31.7 Mb on chromosome 4. In addition to the common associations detected in both and subpopulations, many associated loci were unique to one subpopulation. For example, and were specifically associated with panicle length (PL) in and rice, respectively. Moreover, the -mediated flowering genes and were associated with the formation of panicle architecture in rice. These results suggest that different gene networks regulate panicle architecture in and rice.

A Novel Tiller Angle Gene, TAC3, together with TAC1 and D2 Largely Determine the Natural Variation of Tiller Angle in Rice Cultivars.

Tiller angle is one of the most important components of the ideal plant architecture that can greatly enhance rice grain yield. Understanding the genetic basis of tiller angle and mining favorable alleles will be helpful for breeding new plant-type varieties. Here, we performed genome-wide association studies (GWAS) to identify genes controlling tiller angle using 529 diverse accessions of Oryza sativa including 295 indica and 156 japonica accessions in two environments. We identified 7 common quantitative trait loci (QTLs), including the previously reported major gene Tiller Angle Control 1 (TAC1), in the two environments, 10 and 13 unique QTLs in Hainan and Wuhan, respectively. More QTLs were identified in indica than in japonica, and three major QTLs (qTA3, qTA1b/DWARF2 (D2) and qTA9c/TAC1) were fixed in japonica but segregating in indica, which explained the wider variation observed in indica compared with that in japonica. No common QTLs were identified between the indica and japonica subpopulations. Mutant analysis for the candidate gene of qTA3 on chromosome 3 indicated a novel gene, Tiller Angle Control 3 (TAC3), encoding a conserved hypothetical protein controlling tiller angle. TAC3 is preferentially expressed in the tiller base. The ebisu dwarf (d2) mutant exhibited a decreased tiller angle, in addition to its previously described abnormal phenotype. A nucleotide diversity analysis revealed that TAC3, D2 and TAC1 have been subjected to selection during japonica domestication. A haplotype analysis identified favorable alleles of TAC3, D2 and TAC1, which may be used for breeding plants with an ideal architecture. In conclusion, there is a diverse genetic basis for tiller angle between the two subpopulations, and it is the novel gene TAC3 together with TAC1, D2, and other newly identified genes in this study that controls tiller angle in rice cultivars.

Genomewide association analysis for awn length linked to the seed shattering gene qSH1 in rice.

Awn is one of the most important domesticated traits in rice (Oryza sativa). Understanding the genetic basis of awn length is important for grain harvest and production, because long awn length is disadvantageous for both grain harvest and milling. We investigated the awn length of 529 rice cultivars and performed a Genomewide association studies (GWAS) in the indica and japonica subpopulations, and the whole population. In total, we found 17 loci associated with awn length. Of these loci, seven were linked to previously reported quantitative trait loci, and one was linked to the awn gene An-1. Nine novel loci were repeatedly identified in different environments. One of the nine associations was identified in both the whole and japonica populations. Special interest was the detection of the most significant association SNP, sf0136352825, which was less than 95 kb from the seed shattering gene qSH1. These results may provide potentially favourable haplotypes for molecular breeding in rice.

Regulatory role of FZP in the determination of panicle branching and spikelet formation in rice.

FRIZZLE PANICLE (FZP) and RFL/ABERRANT PANICLE ORGANIZATION 2 (APO2) play important roles in regulating the ABCDE floral organ identity genes. However, the relationships among FZP and these floral identity genes in the regulation of panicle formation remain unclear. Here, we used the novel mutant fzp-11, wild-type and FZP-overexpressing plants to compare the expression of these genes during panicle development by real-time PCR and in situ hybridization. The results indicate that FZP is a major negative regulator of RFL/APO2 and determines the transition from panicle branching to spikelet formation. Moreover, overexpression of FZP severely represses axillary meristem formation in both the vegetative and reproductive phases and the outgrowth of secondary branches in panicle. FZP overexpression positively regulates the expression of a subset of the class B genes, AGL6 genes (OsMADS6 and OsMADS17) as well as class E genes (OsMADS1, OsMADS7 and OsMADS8) in floral meristem (FM). Thus, it suggested that FZP could specify floral organ identity by regulating the related OsMADS-box genes.

Loss of function of OsMADS3 via the insertion of a novel retrotransposon leads to recessive male sterility in rice (Oryza sativa).

Natural mutation is the source of natural variation, which is the fundamental basis for the genetic improvement of crops. During the process of developing a recombinant inbred line (RI), a spontaneous mutagenesis in RI127 led to the production of the recessive male-sterile line RI127S. Via a map-based cloning approach, the gene controlling the male sterility was identified as OsMADS3, which was previously reported to be associated with floral organ development and male sterility. Thermal asymmetric interlaced PCR isolated one 1633-bp insertion in OsMADS3 in RI127S, which damaged its function due to failed transcription. The 1633-bp insertion was derived from a fragment flanked by retrotransposon genes on chromosome 5. Seven haplotypes of OsMADS3 were observed among 529 cultivars and 107 wild rice accessions, and 98% of the investigated genotypes carried the same H2 haplotype, indicating that OsMADS3 is highly conserved. RI127S has the combined genome constitution of its parents, indica rice Teqing and japonica 02428, and carries the widely compatible S5 gene donated by 02428. RI127 exhibits good performance in regard to its agronomic traits and has a wide compatibility. Therefore, RI127S would be an elite mediator for recurrent breeding in cases requiring a tedious hand-crossing-based inter-crossing phase. RI127S can be crossed not only with indica rice but also with japonica rice, thus providing breeders with flexible arrangements in recurrent breeding programs.

Expression patterns of photoperiod and temperature regulated heading date genes in Oryza sativa.

In plants, flowering is a major biological phenomenon, which is regulated by an array of interactions occurring between biotic and abiotic factors. In our study, we have compared the expression profiles of flowering genes involved in the flowering pathway, which are influenced by conditions like photoperiod and temperature from seedling to heading developmental stages in two Oryza sativa indica varieties, viz., Zhenshan 97 and Minghui 63 using a expression network approach. Using the network expression approach, we found 17 co-expressed genes having the same expression profile pattern as three key photoperiod flowering genes Hd1, Ehd1 and Hd3a. We also demonstrated that these three co-expressed genes have a similar simulation pattern as temperature flowering genes. Based on our observations, we hypothesize that photoperiod and temperature regulate flowering pathways independently. The present study provides a basis for understanding the network of co-expressed genes involved in flowering pathway and presents a way to demonstrate the behavior of specific gene sets in specific cultivars.

[Ghd7, a pleiotropic gene controlling flag leaf area in rice].

Photosynthesis is the unique source of energy for plant. Flag leaf contributed the majority of photosynthate after rice flowering. Ghd7 is a pleiotropic gene, which can significantly increase rice production. In order to study the genetic effects of Ghd7 on the flag leaf morphology, we made quantitative trait locus (QTL) analysis for flag leaf length (FLL), flag leaf width (FLW), and flag leaf area (FLA) using a Ghd7-BC2F2 population of 190 plants. In the BC2F2 population, the frequency distribution of FLL, FLW, and FLA were bimodal and in agreement with single Mendelian segregation ratio (3:1). FLL, FLW, and FLA were positively correlated with grains per panicle in the population. One QTL was mapped to the in-terval between markers RM3859 and C39 on chromosome 7, which explained 73.3%, 62.3%, and 71.8% of the variations for FLL, FLW, and FLA, and co-segregated with Ghd7. Two near-isogenic lines of NIL (mh7) and NIL (tq7) were devel-oped using Zhenshan 97 as the recurrent parent and Minghui 63 and Teqing as the donor parent, respectively. Both NILs significantly increased the phenotypic values of FLL, FLW, and FLA as compared with Zhenshan 97. FLL, The values of FLW and FLA for Ghd7 over-expression transgenic plants were 8.9 cm, 0.5 cm, and 17.8 cm2 larger than its recipient Heji-ang 19. These results demonstrated that Ghd7 plays an important role in controlling the flag leaf area in rice.

Yield-related QTLs and their applications in rice genetic improvement.

Grain yield is one of the most important indexes in rice breeding, which is governed by quantitative trait loci (QTLs). Different mapping populations have been used to explore the QTLs controlling yield related traits. Primary populations such as F(2) and recombinant inbred line populations have been widely used to discover QTLs in rice genome-wide, with hundreds of yield-related QTLs detected. Advanced populations such as near isogenic lines (NILs) are efficient to further fine-map and clone target QTLs. NILs for primarily identified QTLs have been proposed and confirmed to be the ideal population for map-based cloning. To date, 20 QTLs directly affecting rice grain yield and its components have been cloned with NIL-F(2) populations, and 14 new grain yield QTLs have been validated in the NILs. The molecular mechanisms of a continuously increasing number of genes are being unveiled, which aids in the understanding of the formation of grain yield. Favorable alleles for rice breeding have been 'mined' from natural cultivars and wild rice by association analysis of known functional genes with target trait performance. Reasonable combination of favorable alleles has the potential to increase grain yield via use of functional marker assisted selection.