Panicle Mutant Research Articles

Identification and Gene Mapping of A White Panicle Mutant wp7 in Rice

Identification and Gene Mapping of A White Panicle Mutant <i>wp7</i> in Rice

Erect panicle architecture contributes to increased rice production through the improvement of canopy structure

The erect panicle architecture has contributed to the improvement of yield in japonica rice breeding, and recent molecular analysis has revealed the mechanisms involved in individual plant yield increases. However, the population structure is more important in rice production compared with individual plant yield. Our study compared the population canopy structure of a curved panicle variety Sasanishiki (WT) and an erect panicle mutant derived from CRISPR/Cas9 gene editing at the DENSE AND ERECT PANICLE 1 (DEP1) locus. The results showed that more light could reach to the leaves under the panicle in the CRISPR-dep1 population compared with the WT. The canopy of the CRISPR-dep1 population exhibited higher temperature and lower humidity compared with the WT after heading. A subsequent survey showed that the CO2 concentration in the CRISPR-dep1 population was significantly lower than that in the WT population from full heading to 15 days after heading. Moreover, the increase of biomass in the CRISPR-dep1 population was greater than that in the WT. We noticed that the CRISPR-dep1 mutant could achieve higher yield under low fertilization application compared with the WT under high fertilizer application through increased transplant density. These traits could contribute to an agricultural sustainable development strategy. The quality investigation showed that the dep1 allele increased the yield along with imposing a penalty on grain quality. Our study not only elucidated the mechanism of yield improvement in an erect panicle architecture variety from the perspective of population structure optimization but also provides a theoretical basis for supporting cultivation systems with the erect panicle architecture.

Generation and Characterization of a Foxtail Millet (Setaria italica) Mutant Library.

Foxtail millet (Setaria italica) is attractive to plant scientists as a model plant because of several distinct characteristics, such as its short stature, rapid life cycle, sufficient seed production per plant, self-compatibility, true diploid nature, high photosynthetic efficiency, small genome size, and tolerance to abiotic and biotic stress. However, the study on the genetic resources of foxtail millet largely lag behind those of the other model plants such as Arabidopsis, rice and maize. Mutagenized populations cannot only create new germplasm resources, but also provide materials for gene function research. In this manuscript, an ethyl methanesulfonate (EMS)-induced foxtail millet population comprising ∼15,000 individual M1 lines was established. Total 1353 independent lines with diverse abnormal phenotypes of leaf color, plant morphologies and panicle shapes were identified in M2. Resequencing of sixteen randomly selected M2 plants showed an average estimated mutation density of 1 loci/213 kb. Moreover, we provided an example for rapid cloning of the WP1 gene by a map-based cloning method. A white panicle mutant, named as wp1.a, exhibited significantly reduced chlorophyll (Chl) and carotenoid contents in leaf and panicle. Map-based cloning results showed an eight-base pair deletion located at the sixth exon of wp1.a in LOC101786849, which caused the premature termination. WP1 encoded phytoene synthase. Moreover, the sequencing analysis and cross test verified that a white panicle mutant wp1.b was an allelic mutant of wp1.a. The filed phenotypic observation and gene cloning example showed that our foxtail millet EMS-induced mutant population would be used as an important resource for functional genomics studies of foxtail millet.

イネ閉花受粉性変異体spw1-clsおよび疎粒変異体M645(lax2-3アレル)識別用PCR-based DNAマーカーの開発

イネ閉花受粉性変異体spw1-clsおよび疎粒変異体M645(lax2-3アレル)識別用PCR-based DNAマーカーの開発

Genetic analysis of dense and erect panicle 2 allele DEP2-1388 and its application in hybrid rice breeding

Using ethyl methanesulfonate (EMS) mutagenesis, we isolated an erect panicle mutant, R1338, from indica heavy-panicle restorer Shuhui498. Compared with wild type control, the mutant displayed dwarfism, erect and short panicle, short primary panicle branch, increased grain density, short grain length and increased grain thickness. In addition, the erect panicle architecture of R1388 resulted in significant decreased bending moment and increased resistance to panicle bending. Histocytological analysis indicated that the diameter of uppermost internode, cellulose content and lignin content play important roles in resistance to panicle bending. Genetic analysis revealed that the mutant phenotype was controlled by a semi-dominant nuclear gene. With resequencing and MutMap analysis strategy, we found that one SNP from A to G at the seventh exon of DEP2 resulted in the 928(th) amino acid substitution from arginine (AGG) to glycine (GGG) in R1338 mutant. Considering the phenotype of other dep2 mutants, the phenotype of R1338 was likely to be caused by the SNP in DEP2. The mutant R1338 and wild type were crossed with several sterile lines which respectively had different panicle types, the combinations generated from R1338 and curve panicle sterile lines showed semi-erect panicle, higher seed setting percentage and heterosis, and the combinations generated from R1388 and erect panicle sterile line with DEP1 showed erect panicle by gene additive effect. Moreover, the combinations with semi-erect panicle had superior light transmittance and stronger light intensity, which improved efficiency of light utilization to intermediate and subjacent leaves compared to the combinations with curved panicle. This study provides a good strategy to solve the problem of population density in three-line hybrid rice.

Genetic Mapping of a Rice Loose Upper Panicle Mutant

We identified a loose upper panicle mutant (lup) from a japonica-type rice variety, Hwacheongbyeo, treated by Ethyl Methane Sulfonate (EMS). The lup mutant displayed an increased distance between spikelets particularly in the first primary branches, and the number of spikelet was reduced. In addition, aborted spikelets in the tip of first primary branches were observed. Besides these morphological changes in the panicle, the lup mutant also displayed overall reduction in culm length, panicle length, grain weight, and tiller number. On the contrary, the chlorophyll content was relatively high in lup mutant in comparison to wild-type plants, and displayed a stay-green phenotype even after physiological maturity. Genetic analysis (using F2 population of lup/M.23) revealed that a single recessive gene is involved in the above-mentioned morphological changes in the lup mutant. A candidate genomic region was fine-mapped at an interval of 1.04 Mb flanked by two molecular markers, 18170 and D0052, on the long arm of chromosome 8. In this region, we found a total of 348 mutation points using a slightly modified MutMap method. Based on these results, we expect the candidate genomic region containing a putative LUP gene will provide an important clue in developmental regulation of spikelets and panicle in rice.

Identification and Gene Fine Mapping of White Panicle Mutant wp4 in Oryza sativa

Identification and Gene Fine Mapping of White Panicle Mutant <i>wp4</i> in <i>Oryza sativa</i>

Morphological characteristics and gene mapping of a dense panicle (dp2) mutant in rice (Oryza sativa L.)

A dense panicle mutant (dp2) derived from the Oryza sativa ssp. japonica cultivar Nipponbare through ethyl methane sulfonate mutagenesis was used in present study. Compared to the wild type, the panicle of dp2 mutant exhibited more branches and denser grains. Further more, the number of spikelets per panicle, number of primary branches and secondary branches of dp2 mutant were significantly increased while the panicle length, and 1,000-grain weight were significantly decreased. The results from the genetic analysis indicated that the dense panicle phenotype was controlled by a single dominance nuclear gene. Polymorphic analysis of SSR and InDel markers demonstrated that the DP2 gene was located at the long arm of chromosome 2, which was further mapped between SSR markers RM341 and RM13356 in a physical region of 398 kb. Within this region, the RCN2 (LOC_Os02g32950) gene which was annotated relating to the development of rice panicle was found. Compared to the wild type, the sequence of RCN2 gene in the dp2 mutant showed that two SNPs replacement had taken place in the promoter region (G–A) and the intron region (A–T), respectively. The dp2 mutant could be a novel mutant of RCN2 gene and this novel mutant might be useful for further studies on this gene.

Genetic Analysis and Mapping of an Enclosed Panicle Mutant Locus esp1 in Rice (Oryza sativa L.)

A mutant was isolated from the M2 of 60Co-γ ray mutagenized male-fertility restorer line Zao-R974 in rice. The mutant showed pleiotropic phenotypes including dwarfism, delayed heading time, short and partially enclosed panicles, short uppermost internode, decreased grain and secondary branch numbers per panicle, and partially degenerated spikelets. The mutant was named as esp1 (enclosed shorter panicle 1). Genetic analysis indicated that the mutant phenotype was controlled by a recessive locus. Spraying exogenous GA3 did not rescue the panicle enclosure. Using an F2 and a BC1 population of the cross between esp1 and a japonica cultivar Nipponbare, we mapped the ESP1 locus to a region of ∼260 kb on chromosome 11. This result provides a basis for further map-based cloning of the ESP1 locus.

Genetic analysis and fine mapping of an enclosed panicle mutant esp2 in rice (Oryza sativa L.)

The phenomenon of panicle enclosure in rice is mainly caused by the shortening of uppermost internode. Elucidating the molecular mechanism of panicle enclosure will be helpful for solving the problem of panicle enclosure in male sterile lines and creating new germplasms in rice. We acquired a monogenic recessive enclosed panicle mutant, named as esp2 (enclosed shorter panicle 2), from the tissue culture progeny of indica rice cultivar Minghui-86. In the mutant, panicles were entirely enclosed by flag leaf sheaths and the uppermost internode was almost completely degenerated, but the other internodes did not have obvious changes in length. Genetic analysis indicated that the mutant phenotype was controlled by a recessive gene, which could be steadily inherited and was not affected by genetic background. Apparently, ESP2 is a key gene for the development of uppermost internode in rice. Using an F2 population of a cross between esp2 and a japonica rice cultivar Xiushui-13 as well as SSR and InDel markers, we fine mapped ESP2 to a 14-kb region on the end of the short arm of chromosome 1. According to the rice genome sequence annotation, only one intact gene exists in this region, namely, a putative phosphatidylserine synthase gene. Sequencing analysis on the mutant and the wild type indicated that this gene was inserted by a 5287-bp retrotransposon sequence. Hence, we took this gene as a candidate of ESP2. The results of this study will facilitate the cloning and functional analysis of ESP2 gene.

Fine mapping and candidate gene analysis of dense and erect panicle 3, DEP3, which confers high grain yield in rice (Oryza sativa L.)

Architecture of the rice inflorescence, which is determined mainly by the morphology, number and length of primary and secondary inflorescence branches, is an important agronomical trait. In the current study, we characterized a novel dense and erect panicle (EP) mutant, dep3, derived from the Oryza sativa ssp. japonica cultivar Hwacheong treated with N-methyl-N-nitrosourea. The panicle of the dep3 mutant remained erect from flowering to full maturation, whereas the panicle of the wild type plant began to droop after flowering. The dep3 mutation also regulated other panicle characteristics, including panicle length, grain shape and grain number per panicle. Anatomical observations revealed that the dep3 mutant had more small vascular bundles and a thicker culm than wild type plants, explaining the EP phenotype. Genetic analysis indicated that the phenotype with the dense and EP was controlled by a single recessive gene, termed dep3. The DEP3 gene was identified as the candidate via a map-based cloning approach and was predicted to encode a patatin-like phospholipase A2 (PLA2) superfamily domain-containing protein. The mutant allele gene carried a 408bp genomic deletion within LOC_Os06g46350, which included the last 47bp coding region of the third exon and the first 361bp of the 3'-untranslated region. Taken together, our results indicated that the patatin-like PLA2 might play a significant role in the formation of vascular bundles, and that the dep3 mutant may provide another EP resource for rice breeding programs.

The screening of mutants and construction of mutant library for Oryza sativa cv. Nipponbare via ethyl methane sulphonate inducing

Following the sequencing of rice genome, the functional analysis of unidentified genes is gaining wide importance. Mutant isolation is one of the effective ways to isolate and clone the target genes and analyze their functions. To find the various mutants in the same genetic background, seeds of Oryza sativa cv. Nipponbare were treated with ethyl methane sulphonate (EMS). A total of 1056 mutants were screened for five categories in M2 generation with the seedling frequency of 26.29‰ at three-leaf stage, but only 264 mutants were verified in M3 generation with a frequency of 6.57‰. Among the mutants verified in M3 generation, the frequency of leaf mutation was the highest (2.22‰), followed by seedling height (1.74‰) and the abiotic stress tolerance mutant (1.47‰). Nineteen characteristic mutations, including a big group of abiotic stress tolerant mutants such as herbicide resistant, salt tolerant and drought tolerant were identified at this stage. By observation of rice growth characteristics at different developmental stages, another 220 mutants have been isolated and verified in the M3 generation with the mutant frequency of 53.9‰ covering about 28 mutant traits. Among those identified, the highest frequencies were obtained for appearance of brown rice mutant with 18.37‰, followed by panicle mutant with 13.47‰, and grain mutants with 9.06‰. All the mutants screened above were suitable for gene function analysis and for utilization in agronomy.

Map-based cloning of the ERECT PANICLE 3 gene in rice

Panicle architecture in rice can have a strong influence on yield. Using N-methyl-N-nitrosourea mutagenesis, we isolated an erect panicle mutant, Hep, from Hwasunchalbyeo, a glutinous japonica rice cultivar. Genetic analysis revealed that the erect panicle phenotype was controlled by a single recessive mutation designated erect panicle 3 (ep3). Genetic mapping revealed that the ep3 mutation was located on the short arm of chromosome 2 in a 0.1 cM region delimited by the STS markers STS5803-5 and STS5803-7. The ep3 locus corresponded to 46.8 kb region and contained six candidate genes. Comparison of the DNA sequences of the candidate genes from wild-type and erect panicle plants revealed a single base-pair change in the second exon of LOC_Os02g15950, which is predicted to result in a nonsense mutation. LOC_Os02g15950 encodes a putative F-box protein containing 515 amino acids and is expressed throughout the plant during all growth stages. A line carrying a T-DNA insertion in LOC_ Os02g15950 was obtained and shown to have the same phenotype as the ep3 mutant, thus confirming the identification of LOC_Os02g15950 as the ERECT PANICLE 3 (EP3) gene. The ep3 mutation causes a significant increase in the number of small vascular bundles as well as the thickness of parenchyma in the peduncle, which results in the erect panicle phenotype.

A Rice Panicle Mutant Created by Transformation with an Antisense cDNA Library

A rice (Oryza sativa L.) mutant displaying defects in panicle development was identified among transformants in a transgenic mutagenized experiment using an antisense cDNA library prepared from young rice panicles. In the mutant, the average spikelet number was reduced to 59.8 compared with 104.3 in wild-type plants. In addition, the seed-setting rate of the mutant was low (39.3%) owing to abnormal female development. Genetic analysis of T1 and T2 progeny showed that the traits segregated in a 3 (mutant): 1 (wild type) ratio and the mutation was cosegregated with the transgene. Southern blot and thermal asymmetric interlaced polymerase chain reaction analyses showed that the mutant had a single T-DNA insertion on chromosome 5, where no gene was tagged. Sequencing analysis found that the transgenic antisense cDNA was derived from a gene encoding an F-box protein in chromosome 7 with unidentified function. This and another four homologous genes encoding putative F-box proteins form a gene cluster. These results indicate that the phenotypic mutations were most likely due to the silencing effect of the expressed transgenic antisense construct on the member(s) of the F-box gene cluster. (Managing editor: Li-Hui Zhao)

Morphological and molecular characterization of a new frizzy panicle mutant, "fzp-9(t)", in rice (Oryza sativa L.)

Morphological and molecular characterization of a new frizzy panicle mutant, "fzp-9(t)", in rice (Oryza sativa L.)

The OsFOR1 gene encodes a polygalacturonase-inhibiting protein (PGIP) that regulates floral organ number in rice.

We have isolated a cDNA clone, OsFOR1, from the immature panicles of rice. The OsFOR1 (Oryza sativa floral organ regulator 1) gene encodes a protein that contains a leucine-rich repeat (LRR) domain. This domain comprises 10 tandem repeats of a canonical 24-amino acid LRR sequence. The structure and the number of LRRs for OsFOR1 are similar to those of polygalacturonase-inhibiting proteins (PGIPs) from various other plant species. Moreover, the OsFOR1 recombinant protein, when fused to maltose-binding protein (MBP), shows PGIP activity against the Aspergillus niger polygalacturonase. OsFOR1 is highly expressed in the calli and immature and mature panicles, while detectable at only low levels in seedling roots and mature stems. In situ hybridization experiments showed the transcripts of OsFOR1 are present in young spikelet primordia and in almost all of the young floral organs. Transgenic approaches were used to study in vivo functioning. Antisense expression of OsFOR1 resulted in an increase in the numbers of floral organs, including the stamen, carpel, palea/lemma, stigma, and lodicule. OsFOR1 transcript was not detected in the frizzy panicle mutant, which is defective in its spikelet formation but normal in inflorescence-meristem initiation and maintenance. Therefore, we suggest that OsFOR1 plays a role in the formation and/or maintenance of floral organ primordia.

FRIZZY PANICLE is required to prevent the formation of axillary meristems and to establish floral meristem identity in rice spikelets.

Inflorescences of grass species have a distinct morphology in which florets are grouped in compact branches called spikelets. Although many studies have shown that the molecular and genetic mechanisms that control floret organ formation are conserved between monocots and dicots, little is known about the genetic pathway leading to spikelet formation. In the frizzy panicle (fzp) mutant of rice, the formation of florets is replaced by sequential rounds of branching. Detailed analyses revealed that several rudimentary glumes are formed in each ectopic branch, indicating that meristems acquire spikelet identity. However, instead of proceeding to floret formation, axillary meristems are formed in the axils of rudimentary glumes and they either arrest or develop into branches of higher order. The fzp mutant phenotype suggests that FZP is required to prevent the formation of axillary meristems within the spikelet meristem and permit the subsequent establishment of floral meristem identity. The FZP gene was isolated by transposon tagging. FZP encodes an ERF transcription factor and is the rice ortholog of the maize BD1 gene. Consistent with observations from phenotypic analyses, FZP expression was found to be restricted to the time of rudimentary glumes differentiation in a half-ring domain at the base of which the rudimentary glume primordium emerged.

Characterization and mapping of a white panicle mutant gene in rice

A spontaneous white panicle mutant was found from the F6 progenies of an indica/japonica cross. The mutant exhibits white stripes on its basal leaves while the panicles, rachis and pedicel are milky white colored at flowering stage. Genetic analysis in an F2 population from the cross of Zhi7/white panicle mutant indicates that the white panicle phenotype is controlled by a single recessive nuclear gene, tentatively termed aswp(t). Using microsatellite markers, thewp(t) gene was anchored between the markers of SSR101 and SSR63.9 with a map distance of 2.3 and 0.8 cM, respectively, and co-segregated with the marker of SSR17 on rice chromosome 1.

Characterization and mapping of a white panicle mutant gene in rice

Characterization and mapping of a white panicle mutant gene in rice

Genetical Studies on Dense and Lax Panicles in Rice : II Character Expression and Mode of Inheritance of Dense Panicle Rice

Following the previous paper (1979) dealt with the lax panicle rice, this investiga-tion was conducted to get information on the character expression of dense panicle rice and its mode of inheritance. It was found that the dense panicle cultivars and strains, in general, possessed either greater number of spikelets and panicle branches, or shorter panicle and primary branches than the normal type cultivar, the, causes for dense panicle being largely varied with the cultivars and strains tested. In some cultivars and strains which had extrerne dense panicles, the occurrence of tertiary branches was observed. It was verified that the newly devised method, that is, the ratio of total spikelets to total length of rachis and primary branches, by which the panicle density of normal and lax panicle rices would be satisfactorily esti.mated, as described in the previous paper (1979), could also be applied to the dense panicle rices. Crosses between a dense panicle mutant strain and its original cultivar. Akibare, and between the former and a dwarf mutant strain induced in Akibare were mad, e, and the segregations of panicle density and its related characters were investigated in F2. It was shown that the dense panicle mutant strain carried a single recessive gene respon-sible for the dense panicle, which was tentatively designated as dn. Positive gene interaction was also observed between the dense panicle gene and the dwarf gene