Fine mapping of the BnaC04.BIL1 gene controlling plant height in Brassica napus L

Background Plant height is an important architecture trait which is a fundamental yield-determining trait in crops. Variety with dwarf or semi-dwarf phenotype is a major objective in the breeding because dwarfing architecture can help to increase harvest index, increase planting density, enhance lodging resistance, and thus be suitable for mechanization harvest. Although some germplasm or genes associated with dwarfing plant type have been carried out. The molecular mechanisms underlying dwarfism in oilseed rape (Brassica napus L.) are poorly understood, restricting the progress of breeding dwarf varieties in this species. Here, we report a new dwarf mutant Bndwarf2 from our B. napus germplasm. We studied its inheritance and mapped the dwarf locus BnDWARF2. Results The inheritance analysis showed that the dwarfism phenotype was controlled by one semi-dominant gene, which was mapped in an interval of 787.88 kb on the C04 chromosome of B. napus by Illumina Brassica 60 K Bead Chip Array. To fine-map BnDWARF2, 318 simple sequence repeat (SSR) primers were designed to uniformly cover the mapping interval. Among them, 15 polymorphic primers that narrowed down the BnDWARF2 locus to 34.62 kb were detected using a F2:3 family population with 889 individuals. Protein sequence analysis showed that only BnaC04.BIL1 (BnaC04g41660D) had two amino acid residues substitutions (Thr187Ser and Gln399His) between ZS11 and Bndwarf2, which encoding a GLYCOGEN SYNTHASE KINASE 3 (GSK3-like). The quantitative real-time PCR (qRT-PCR) analysis showed that the BnaC04.BIL1 gene expressed in all tissues of oilseed rape. Subcellular localization experiment showed that BnaC04.BIL1 was localized in the nucleus in tobacco leaf cells. Genetic transformation experiments confirmed that the BnaC04.BIL1 is responsible for the plant dwarf phenotype in the Bndwarf2 mutants. Overexpression of BnaC04.BIL1 reduced plant height, but also resulted in compact plant architecture. Conclusions A dominant dwarfing gene, BnaC04.BIL1, encodes an GSK3-like that negatively regulates plant height, was mapped and isolated. Our identification of a distinct gene locus may help to improve lodging resistance in oilseed rape.

Position Validation of the Dwarfing Gene Dw6 in Oat (Avena sativa L.) and Its Correlated Effects on Agronomic Traits

An F6:8 recombinant inbred line (RIL) population derived from the cross between WAOAT2132 (Dw6) and Caracas along with the two parents were used to evaluate the genetic effects of Dw6 dwarfing gene on plant height and other agronomic traits in oat (Avena sativa L.) across three environments, and develop closely linked markers for marker-assisted selection (MAS) for Dw6. The two parents differed in all investigated agronomic traits except for the number of whorls. The RIL lines showed a bimodal distribution for plant height in all three tested environments, supporting the height of this population was controlled by a single gene. Dw6 significantly reduced plant height (37.66∼44.29%) and panicle length (13.99∼22.10%) but without compromising the coleoptile length which was often positively associated with the reduced stature caused by dwarfing genes. Dw6 has also strong negative effects on hundred kernel weight (14.00∼29.55%), and kernel length (4.21∼9.47%), whereas the effects of Dw6 on the kernel width were not uniform across three environments. By contrast, lines with Dw6 produced more productive tillers (10.11∼10.53%) than lines without Dw6. All these together suggested the potential yield penalty associated with Dw6 might be partially due to the decrease of kernel weight which is attributed largely to the reduction of kernel length. Eighty-one simple sequence repeat (SSR) primer pairs from chromosome 6D were tested, five of them were polymorphic in two parents and in two contrasting bulks, confirming the 6D location of Dw6. By using the five polymorphic markers, Dw6 was mapped to an interval of 1.0 cM flanked by markers SSR83 and SSR120. Caution should be applied in using this information since maker order conflicts were observed. The close linkages of these two markers to Dw6 were further validated in a range of oat lines. The newly developed markers will provide a solid basis for future efforts both in the identification of Dw6 in oat germplasm and in the determination of the nature of the gene through positional cloning.

A Dwarfing Gene sd1-d (Dee-geo-woo-gen dwarf) on Lodging Resistance and Related Traits in Rice

A dwarfing allele at the sd1 locus on chromosome 1 in rice, sd1-d, has been playing important role for developing lodging-resistant and high-yielding indica varieties IR8 and IR36. The dominant allele SD1 for long culm at the locus is differentiated into SD1-in and SD1-ja that are harbored in indica and japonica subspecies, respectively. The sd1-d of IR36 was substituted with SD1-in or SD1-ja by 17 backcrosses with IR36, and two isogenic tall lines were developed by using an indica variety IR5867 and a japonica one ‘Koshihikari’ as donors, which were denoted by “5867-36” and “Koshi-36’’, respectively. The present study was conducted to examine the effect of dwarfing gene sd1-d on lodging resistance and related traits, compared with SD1-in and SD1-ja. Two isogenic lines and IR36 were cultivated in the field of the Faculty of Agriculture and Marine Science, Kochi University, Japan during 2017. Regarding index of lodging (g·cm/g × 100), genotypes were in the order: 5867-36 (97.4) > Koshi-36 (74.1) > IR36 (46.0) on the 21st-day after 80%-heading, and they were in the same order on 10th-day after 80%-heading. The 4th-panicle length (cm) was in the order: 5867-36 (118.7) > Koshi-36 (97.6) > IR36 (78.6). Similarly, the 4th-top weight (g) was in the order: 5867-36 (12.2) > Koshi-36 (10.2) > IR36 (9.6). The highest breaking strength (g) was recorded in IR36 (1649) followed by 5867-36 (1493) whereas the lowest breaking strength (g) was recorded in Koshi-36 (1360). Consequently, it is inferred that sd1-d enhances lodging resistance due to the decreases in the length and weight above the 4th-internode as well as the increase of breaking strength. The effect of SD1-in on lodging resistance is lower than that of SD1-ja.

Comparative transcriptomic analysis reveals the regulatory mechanism of the gibberellic acid pathway of Tartary buckwheat (Fagopyrum tataricum (L.) Gaertn.) dwarf mutants

BackgroundTartary buckwheat is an important minor crop species with high nutritional and medicinal value and is widely planted worldwide. Cultivated Tartary buckwheat plants are tall and have hollow stems that lodge easily, which severely affects their yield and hinders the development of the Tartary buckwheat industry.MethodsHeifeng No. 1 seeds were treated with ethylmethanesulfonate (EMS) to generate a mutant library. The dwarf mutantftdmwas selected from the mutagenized population, and the agronomic characteristics giving rise to the dwarf phenotype were evaluated. Ultra-fast liquid chromatography-electrospray ionization tandem mass spectrometry (UFLC-ESI–MS/MS) was performed to determine the factors underlying the different phenotypes between the wild-type (WT) andftdmplants. In addition, RNA sequencing (RNA-seq) was performed via the HiSeq 2000 platform, and the resulting transcriptomic data were analysed to identify differentially expressed genes (DEGs). Single-nucleotide polymorphism (SNP) variant analysis revealed possible sites associated with dwarfism. The expression levels of the potential DEGs between the WT andftdmmutant were then measured via qRT-PCR and fragments per kilobase of transcript per million mapped reads (FPKM).ResultThe plant height (PH) of theftdmmutant decreased to 42% of that of the WT, and compared with the WT, the mutant and had a higher breaking force (BF) and lower lodging index (LI). Lower GA4 and GA7 contents and higher contents of jasmonic acid (JA), salicylic acid (SA) and brassinolactone (BR) were detected in the stems of theftdmmutant compared with the WT. Exogenous application of GAs could not revert the dwarfism of theftdmmutant. On the basis of the transcriptomic analysis, 146 homozygous SNP loci were identified. In total, 12 DEGs with nonsynonymous mutations were ultimately identified, which were considered potential candidate genes related to the dwarf trait. When the sequences of eight genes whose expression was downregulated and four genes whose expression was upregulated were compared, SKIP14, an F-box protein whose sequence is 85% homologous to that of SLY1 in Arabidopsis, presented an amino acid change (from Ser to Asn) and was expressed at a lower level in the stems of theftdmmutant compared with the WT. Hence, we speculated that this amino acid change in SKIP14 resulted in a disruption in GA signal transduction, indirectly decreasing the GA content and downregulating the expression of genes involved in GA biosynthesis or the GA response. Further studies are needed to determine the molecular basis underlying the dwarf phenotype of theftdmmutant.ConclusionWe report a Tartary buckwheat EMS dwarf mutant,ftdm, suitable for high-density planting and commercial farming. A significant decrease in GA4 and GA7 levels was detected in theftdmmutant, and 12 DEGs expressed in the stems of theftdmmutant were selected as candidates of the dwarfing gene. One nonsynonymous mutation was detected in theSKIP14gene in theftdmmutant, and this gene had a lower transcript level compared with that in the WT.

The semi-dwarfing gene Rht-dp from dwarf polish wheat (Triticum polonicum L.) is the "Green Revolution” gene Rht-B1b

Background The wheat dwarfing gene increases lodging resistance, the grain number per spike and harvest index. Dwarf Polish wheat (Triticum polonicum L., 2n = 4x = 28, AABB, DPW), initially collected from Tulufan, Xinjiang, China, carries a semi-dwarfing gene Rht-dp on chromosome 4BS. However, Rht-dp and its dwarfing mechanism are unknown. Results Homologous cloning and mapping revealed that Rht-dp is the ‘Green Revolution’ gene Rht-B1b. A haplotype analysis in 59 tetraploid wheat accessions showed that Rht-B1b was only present in T. polonicum. Transcriptomic analysis of two pairs of near-isogenic lines (NILs) of DPW × Tall Polish wheat (Triticum polonicum L., 2n = 4x = 28, AABB, TPW) revealed 41 differentially expressed genes (DEGs) as potential dwarfism-related genes. Among them, 28 functionally annotated DEGs were classed into five sub-groups: hormone-related signalling transduction genes, transcription factor genes, cell wall structure-related genes, reactive oxygen-related genes, and nitrogen regulation-related genes. Conclusions These results indicated that Rht-dp is Rht-B1b, which regulates pathways related to hormones, reactive oxygen species, and nitrogen assimilation to modify the cell wall structure, and then limits cell wall loosening and inhibits cell elongation, thereby causing dwarfism in DPW.

The semi-dwarfing gene Rht-dp from dwarf Polish wheat (Triticum polonicum L.) is the “Green Revolution” gene Rht-B1b

Background: The wheat dwarfing gene increases lodging resistance, the grain number per spike and harvest index. Dwarf Polish wheat (Triticum polonicum L., 2n = 4x = 28, AABB, DPW), initially collected from Tulufan, Xinjiang, China, carries a semi-dwarfing gene Rht-dp on chromosome 4BS. However, Rht-dp and its dwarfing mechanism are unknown.Results: Homologous cloning and mapping revealed that Rht-dp is the ‘Green Revolution’ gene Rht-B1b. A haplotype analysis in 59 tetraploid wheat accessions showed that Rht-B1b was only present in T. polonicum. Transcriptomic analysis of two pairs of near-isogenic lines (NILs) of DPW×Tall Polish wheat (Triticum polonicum L., 2n = 4x = 28, AABB, TPW) revealed 41 differentially expressed genes (DEGs) as potential dwarfism-related genes. Among them, 28 functionally annotated DEGs were classed into five sub-groups: hormone-related signalling transduction genes, transcription factor genes, cell wall structure-related genes, reactive oxygen-related genes, and nitrogen regulation-related genes. Conclusions: These results indicated that Rht-dp is Rht-B1b, which regulates pathways related to hormones, reactive oxygen species, and nitrogen assimilation to modify the cell wall structure, and then limits cell wall loosening and inhibits cell elongation, thereby causing dwarfism in DPW.

The semi-dwarfing gene Rht-dp from dwarf Polish wheat (Triticum polonicum L.) is the “Green Revolution” gene Rht-B1b

Background: The wheat dwarfing gene increases lodging resistance, the grain number per spike and harvest index. Dwarf Polish wheat (Triticum polonicum L., 2n = 4x = 28, AABB, DPW), initially collected from Tulufan, Xinjiang, China, carries a semi-dwarfing gene Rht-dp on chromosome 4BS. However, Rht-dp and its dwarfing mechanism are unknown.Results: Homologous cloning and fine mapping revealed that Rht-dp is the ‘Green Revolution’ gene Rht-B1b. A haplotype analysis in 59 tetraploid wheat accessions showed that Rht-B1b was only present in T. polonicum. Transcriptomic analysis of two pairs of near-isogenic lines (NILs) of DPW×Tall Polish wheat (Triticum polonicum L., 2n = 4x = 28, AABB, TPW) revealed 41 differentially expressed genes (DEGs) as potential dwarfism-related genes. Among them, 28 functionally annotated DEGs were classed into five sub-groups: hormone-related signalling transduction genes, transcription factor genes, cell wall structure-related genes, reactive oxygen-related genes, and nitrogen regulation-related genes. Conclusions: These results indicated that Rht-dp is Rht-B1b, which regulates pathways related to hormones, reactive oxygen species, and nitrogen assimilation to modify the cell wall structure, and then limits cell wall loosening and inhibits cell elongation, thereby causing dwarfism in DPW.

Phenology and Dwarfing Gene Interaction Effects on the Adaptation of Selected Wheat (Triticum aestivum L.) Advanced Lines across Diverse Water-Limited Environments of Western Australia

Photoperiod, vernalization, and plant height controlling genes are major developmental genes in wheat that govern environmental adaptation and hence, knowledge on the interaction effects among different alleles of these genes is crucial in breeding cultivars for target environments. The interaction effects among these genes were studied in nineteen Australian advanced lines from diverse germplasm pools and four commercial checks. Diagnostic markers for the Vrn-A1 locus revealed the presence of the spring allele Vrn-A1a in 10 lines and Vrn-A1c in one line. The dominant alleles of Vrn-B1a and Vrn-D1a were identified in 19 and 8 lines, respectively. The most common photoperiod-insensitive allele of Ppd-D1a was identified in 19 lines and three and four copy photoperiod-insensitive alleles (Ppd-B1a and Ppd-B1c) were present in five and one lines, respectively. All the lines were photoperiod-sensitive for the Ppd-A1 locus. All lines were semi-dwarf, having either of the two dwarfing alleles; 14 lines had the Rht-B1b (Rht-1) and the remaining had the Rht-D1b (Rht-2) dwarfing allele. The presence of the photoperiod-insensitive allele Ppd-D1a along with one or two spring alleles at the Vrn1 loci resulted in an earlier heading and better yield. Dwarfing genes were found to modify the heading time—the Rht-D1b allele advanced heading by three days and also showed superior effects on yield-contributing traits, indicating its beneficial role in yield under rain-fed conditions along with an appropriate combination of photoperiod and vernalization alleles. This study also identified the adaptability value of these allelic combinations for higher grain yield and protein content across the different the water-limited environments.

Overgrowth mutants determine the causal role of gibberellin GA2oxidaseA13 in Rht12 dwarfism of wheat

The induced dwarf mutant Rht12 was previously shown to have agronomic potential to replace the conventional DELLA mutants Rht-B1b/Rht-D1b in wheat. The Rht12 dwarfing gene is not associated with reduced coleoptile length (unlike the DELLA mutants) and it is dominant, characteristics which are shared with the previously characterized dwarfing genes Rht18 and Rht14. Using the Rht18/Rht14 model, a gibberellin (GA) 2-oxidase gene was identified in the Rht12 region on chromosome 5A. A screen for suppressor mutants in the Rht12 background identified tall overgrowth individuals that were shown to contain loss-of-function mutations in GA2oxidaseA13, demonstrating the role of this gene in the Rht12 dwarf phenotype. It was concluded that Rht12, Rht18, and Rht14 share the same height-reducing mechanism through the increased expression of GA 2-oxidase genes. Some of the overgrowth mutants generated in this study were semi-dwarf and taller than the original Rht12 dwarf, providing breeders with new sources of agronomically useful dwarfism.

The semi-dwarfing gene Rht-dp from dwarf Polish wheat (Triticum polonicum L.) is the “Green Revolution” gene Rht-B1b

Background The wheat dwarfing gene improves lodging resistance, and increases the grain number per spike and harvest index. Dwarf polish wheat (Triticum polonicum L., 2n = 4x = 28, AABB, DPW), initially collected from Tulufan, Xinjiang, China, carries a semi-dwarfing gene Rht-dp on chromosome 4BS. However, Rht-dp and its dwarfing mechanism are unknown. Results Homologous cloning and fine mapping revealed that Rht-dp is the ‘Green Revolution’ gene Rht-B1b. A haplotype analysis showed that Rht-B1b was only present in T. polonicum. Transcriptomic analysis of two pairs of near-isogenic lines (NILs) of DPW × TPW revealed 41 differentially expressed genes (DEGs) as potential dwarfism-related genes. Among them, 28 functionally annotated DEGs were classed into five sub-groups: hormone-related signalling transduction genes, transcription factor genes, cell wall structure-related genes, reactive oxygen-related genes, and nitrogen regulation-related genes. Conclusions These results indicated that Rht-dp is Rht-B1b, which regulates pathways related to hormones, reactive oxygen species, and nitrogen assimilation to modify the cell wall structure, and then limits cell wall loosening and inhibits cell elongation, thereby causing dwarfism in DPW.