Research

19 pages, 4806 KiB

Open AccessArticle

Development of Single-Segment Substitution Lines and Fine-Mapping of qSPP4 for Spikelets Per Panicle and qGW9 for Grain Width Based on Rice Dual-Segment Substitution Line Z783

by Keli Deng, Han Zhang, Jiayi Wu, Zhuowen Zhao, Dachuang Wang, Guangyi Xu, Jinjin Yu, Yinghua Ling and Fangming Zhao

Int. J. Mol. Sci. 2023, 24(24), 17305; https://doi.org/10.3390/ijms242417305 - 09 Dec 2023

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Abstract

Single segment substitution line (SSSL) libraries are an ideal platform for breeding by design. To develop SSSLs-Xihui18 covering the whole genome, a novel rice chromosome segment substitution line (CSSL), Z783, carrying two substitution segments (average length of 6.55 Mb) on Chr.4 and Chr.9 [...] Read more.

Single segment substitution line (SSSL) libraries are an ideal platform for breeding by design. To develop SSSLs-Xihui18 covering the whole genome, a novel rice chromosome segment substitution line (CSSL), Z783, carrying two substitution segments (average length of 6.55 Mb) on Chr.4 and Chr.9 was identified, which was a gap in the library previously. Z783 was developed from the progeny of recipient “Xihui18” (an indica restorer line) and donor “Huhan3” (a japonica cultivar) by advanced backcross combined molecular marker-assisted selection (MAS). It displayed multiple panicles and less spikelets and wide grains. Then, a F₂ population derived from Xihui18/Z783 was used to map quantitative trait loci (QTLs) for yield-related traits by the mixed linear model method. Nine QTLs were detected (p < 0.05). Furthermore, three SSSLs were constructed by MAS, and all 9 QTLs could be validated, and 15 novel QTLs could be detected by these SSSLs by a one-way ANOVA analysis. The genetic analysis showed that qSSP4 for less spikelets and qGW9 for wide grain all displayed dominant gene action in their SSSLs. Finally, qSSP4 and qGW9 were fine-mapped to intervals of 2.75 Mb and 1.84 Mb, on Chromosomes 4 and 9, respectively. The results lay a solid foundation for their map cloning and molecular breeding by design. Full article

(This article belongs to the Special Issue Rice Molecular Breeding and Genetics 2.0)

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22 pages, 5138 KiB

Open AccessArticle

Down-Regulation of Rice Glutelin by CRISPR-Cas9 Gene Editing Decreases Carbohydrate Content and Grain Weight and Modulates Synthesis of Seed Storage Proteins during Seed Maturation

by Deepanwita Chandra, Kyoungwon Cho, Hue Anh Pham, Jong-Yeol Lee and Oksoo Han

Int. J. Mol. Sci. 2023, 24(23), 16941; https://doi.org/10.3390/ijms242316941 - 29 Nov 2023

Viewed by 1366

Abstract

The glutelins are a family of abundant plant proteins comprised of four glutelin subfamilies (GluA, GluB, GluC, and GluD) encoded by 15 genes. In this study, expression of subsets of rice glutelins were suppressed using CRISPR-Cas9 gene-editing technology to generate three transgenic rice [...] Read more.

The glutelins are a family of abundant plant proteins comprised of four glutelin subfamilies (GluA, GluB, GluC, and GluD) encoded by 15 genes. In this study, expression of subsets of rice glutelins were suppressed using CRISPR-Cas9 gene-editing technology to generate three transgenic rice variant lines, GluA1, GluB2, and GluC1. Suppression of the targeted glutelin genes was confirmed by SDS-PAGE, Western blot, and q-RT-PCR. Transgenic rice variants GluA1, GluB2, and GluC1 showed reduced amylose and starch content, increased prolamine content, reduced grain weight, and irregularly shaped protein aggregates/protein bodies in mature seeds. Targeted transcriptional profiling of immature seeds was performed with a focus on genes associated with grain quality, starch content, and grain weight, and the results were analyzed using the Pearson correlation test (requiring correlation coefficient absolute value ≥ 0.7 for significance). Significantly up- or down-regulated genes were associated with gene ontology (GO) and KEGG pathway functional annotations related to RNA processing (spliceosomal RNAs, group II catalytic introns, small nucleolar RNAs, microRNAs), as well as protein translation (transfer RNA, ribosomal RNA and other ribosome and translation factors). These results suggest that rice glutelin genes may interact during seed development with genes that regulate synthesis of starch and seed storage proteins and modulate their expression via post-transcriptional and translational mechanisms. Full article

(This article belongs to the Special Issue Rice Molecular Breeding and Genetics 2.0)

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16 pages, 4060 KiB

Open AccessArticle

Quantitative Trait Loci Mapping Identified Candidate Genes Involved in Plant Height Regulation in Rice

by Jae-Ryoung Park, Yoon-Hee Jang, Eun-Gyeong Kim, Sang-Sun Hur and Kyung-Min Kim

Int. J. Mol. Sci. 2023, 24(23), 16895; https://doi.org/10.3390/ijms242316895 - 29 Nov 2023

Cited by 1 | Viewed by 655

Abstract

Rice plant height is an agricultural trait closely related to biomass, lodging tolerance, and yield. Identifying quantitative trait loci (QTL) regions related to plant height regulation and developing strategies to screen potential candidate genes can improve agricultural traits in rice. In this study, [...] Read more.

Rice plant height is an agricultural trait closely related to biomass, lodging tolerance, and yield. Identifying quantitative trait loci (QTL) regions related to plant height regulation and developing strategies to screen potential candidate genes can improve agricultural traits in rice. In this study, a double haploid population (CNDH), derived by crossing ‘Cheongcheong’ and ‘Nagdong’ individuals, was used, and a genetic map was constructed with 222 single-sequence repeat markers. In the RM3482-RM212 region on chromosome 1, qPh1, qPh1-1, qPh1-3, qPh1-5, and qPh1-6 were identified for five consecutive years. The phenotypic variance explained ranged from 9.3% to 13.1%, and the LOD score ranged between 3.6 and 17.6. OsPHq1, a candidate gene related to plant height regulation, was screened in RM3482-RM212. OsPHq1 is an ortholog of gibberellin 20 oxidase 2, and its haplotype was distinguished by nine SNPs. Plants were divided into two groups based on their height, and tall and short plants were distinguished and clustered according to the expression level of OsPHq1. QTLs and candidate genes related to plant height regulation, and thus, biomass regulation, were screened and identified in this study, but the molecular mechanism of the regulation remains poorly known. The information obtained in this study will help develop molecular markers for marker-assisted selection and breeding through rice plant height control. Full article

(This article belongs to the Special Issue Rice Molecular Breeding and Genetics 2.0)

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18 pages, 4253 KiB

Open AccessArticle

Osa-miR11117 Targets OsPAO4 to Regulate Rice Immunity against the Blast Fungus Magnaporthe oryzae

by Shang Gao, Yan Hou, Qiwei Huang, Pengzhi Wu, Zhikai Han, Danhong Wei, Huabin Xie, Fengwei Gu, Chun Chen and Jiafeng Wang

Int. J. Mol. Sci. 2023, 24(22), 16052; https://doi.org/10.3390/ijms242216052 - 07 Nov 2023

Cited by 1 | Viewed by 895

Abstract

The intricate regulatory process governing rice immunity against the blast fungus Magnaporthe oryzae remains a central focus in plant–pathogen interactions. In this study, we investigated the important role of Osa-miR11117, an intergenic microRNA, in regulating rice defense mechanisms. Stem-loop qRT-PCR analysis showed that [...] Read more.

The intricate regulatory process governing rice immunity against the blast fungus Magnaporthe oryzae remains a central focus in plant–pathogen interactions. In this study, we investigated the important role of Osa-miR11117, an intergenic microRNA, in regulating rice defense mechanisms. Stem-loop qRT-PCR analysis showed that Osa-miR11117 is responsive to M. oryzae infection, and overexpression of Osa-miR11117 compromises blast resistance. Green fluorescent protein (GFP)-based reporter assay indicated OsPAO4 is one direct target of Osa-miR11117. Furthermore, qRT-PCR analysis showed that OsPAO4 reacts to M. oryzae infection and polyamine (PA) treatment. In addition, OsPAO4 regulates rice resistance to M. oryzae through the regulation of PA accumulation and the expression of the ethylene (ETH) signaling genes. Taken together, these results suggest that Osa-miR11117 is targeting OsPAO4 to regulate blast resistance by adjusting PA metabolism and ETH signaling pathways. Full article

(This article belongs to the Special Issue Rice Molecular Breeding and Genetics 2.0)

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15 pages, 4606 KiB

Open AccessArticle

Improvement of Rice Blast Resistance in TGMS Line HD9802S through Optimized Anther Culture and Molecular Marker-Assisted Selection

by Pingli Chen, Guanjun Gao, Guangming Lou, Jie Hu, Yufu Wang, Rongjia Liu, Da Zhao, Qing Liu, Bingrui Sun, Xingxue Mao, Liqun Jiang, Jing Zhang, Shuwei Lv, Hang Yu, Wenfeng Chen, Zhilan Fan, Chen Li and Yuqing He

Int. J. Mol. Sci. 2023, 24(19), 14446; https://doi.org/10.3390/ijms241914446 - 22 Sep 2023

Cited by 1 | Viewed by 864

Abstract

Rice blast caused by Magnaporthe oryzae is one of the most serious rice diseases worldwide. The early indica rice thermosensitive genic male sterile (TGMS) line HD9802S has the characteristics of stable fertility, reproducibility, a high outcrossing rate, excellent rice quality, and strong combining [...] Read more.

Rice blast caused by Magnaporthe oryzae is one of the most serious rice diseases worldwide. The early indica rice thermosensitive genic male sterile (TGMS) line HD9802S has the characteristics of stable fertility, reproducibility, a high outcrossing rate, excellent rice quality, and strong combining ability. However, this line exhibits poor blast resistance and is highly susceptible to leaf and neck blasts. In this study, backcross introduction, molecular marker-assisted selection, gene chipping, anther culture, and resistance identification in the field were used to introduce the broad-spectrum blast-resistance gene R6 into HD9802S to improve its rice blast resistance. Six induction media were prepared by varying the content of each component in the culture medium. Murashige and Skoog’s medium with 3 mg/L 2,4-dichlorophenoxyacetic acid, 2 mg/L 1-naphthaleneacetic acid, and 1 mg/L kinetin and N6 medium with 800 mg/L casein hydrolysate, 600 mg/L proline, and 500 mg/L glutamine could improve the callus induction rate and have a higher green seedling rate and a lower white seedling rate. Compared to HD9802S, two doubled haploid lines containing R6 with stable fertility showed significantly enhanced resistance to rice blast and no significant difference in spikelet number per panicle, 1000-grain weight, or grain shape. Our findings highlight a rapid and effective method for improving rice blast resistance in TGMS lines. Full article

(This article belongs to the Special Issue Rice Molecular Breeding and Genetics 2.0)

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14 pages, 1605 KiB

Open AccessArticle

Combining Ability Analysis of Yield-Related Traits of Two Elite Rice Restorer Lines in Chinese Hybrid Rice

by Shiguang Wang, Haoxiang Wu, Zhanhua Lu, Wei Liu, Xiaofei Wang, Zhiqiang Fang and Xiuying He

Int. J. Mol. Sci. 2023, 24(15), 12395; https://doi.org/10.3390/ijms241512395 - 03 Aug 2023

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Abstract

Hybrid rice breeding is an important strategy for enhancing grain yield. Breeding high-performance parental lines and identifying combining abilities is a top priority for hybrid breeding. Yuenongsimiao (YNSM) and its derivative variety Yuehesimiao (YHSM) are elite restorer lines with a high ability of [...] Read more.

Hybrid rice breeding is an important strategy for enhancing grain yield. Breeding high-performance parental lines and identifying combining abilities is a top priority for hybrid breeding. Yuenongsimiao (YNSM) and its derivative variety Yuehesimiao (YHSM) are elite restorer lines with a high ability of fertility restoration, from which 67 derived hybrid combinations have been authorized to different degrees in more than 110 instances in China. In this study, we found that YNSM and YHSM contained three candidate restorer-of-fertility (Rf) genes, Rf3, Rf4, and Rf5/Rf1a, that might confer their restoration ability. Subsequently, we investigated heterosis and combining ability of YNSM and YHSM using 50 F₁ hybrids from a 5 × 10 incomplete diallelic mating design. Our results indicated that hybrid combinations exhibited significant genetic differences, and the additive effects of the parental genes played a preponderant role in the inheritance of observed traits. The metrics of plant height (PH), 1000-grain weight (TGW), panicle length (PL), and the number of spikelets per panicle (NSP) were mainly affected by genetic inheritance with higher heritability. Notably, the general combining ability (GCA) of YHSM exhibited the largest positive effect on the number of grains per panicle (NGP), NSP, PL, and TGW. Thus, YHSM had the largest GCA effect on yield per plant (YPP). In addition, the GCA of YNSM exhibited a positive impact on YPP, mainly due to the critical contribution of seed setting percentage (SSP). Moreover, YNSM and YHSM exhibited negative GCA effects on PH, implying that YNSM and YHSM could effectively enhance plant lodging resistance by reducing the plant height of the derived hybrids. Remarkably, among the hybrids, Yuanxiang A/YNSM (YXA/YNSM), Shen 08S/Yuemeizhan (S08S/YMZ), and Quan 9311A/YHSM (Q9311A/YHSM) represent promising new combinations with a higher specific combining ability (SCA) effect value on YPP with a value more than 3.50. Our research thus highlights the promising application for the rational utilization of YNSM and YHSM in hybrid rice breeding. Full article

(This article belongs to the Special Issue Rice Molecular Breeding and Genetics 2.0)

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16 pages, 5502 KiB

Open AccessArticle

Transcriptome Analyses Show Changes in Gene Expression Triggered by a 31-bp InDel within OsSUT3 5′UTR in Rice Panicle

by Qiuping Li, Chunlong Zhang, Jiancheng Wen, Lijuan Chen, Yitong Shi, Qinghui Yang and Dandan Li

Int. J. Mol. Sci. 2023, 24(13), 10640; https://doi.org/10.3390/ijms241310640 - 26 Jun 2023

Viewed by 911

Abstract

Pollen development and its fertility are obligatory conditions for the reproductive success of flowing plants. Sucrose transporter 3 (OsSUT3) is known to be preferentially expressed and may play critical role in developing pollen. A 31-bp InDel was identified as a unique [...] Read more.

Pollen development and its fertility are obligatory conditions for the reproductive success of flowing plants. Sucrose transporter 3 (OsSUT3) is known to be preferentially expressed and may play critical role in developing pollen. A 31-bp InDel was identified as a unique variation and was shown to be responsible for the expression of downstream gene in our previous study. In this study, to analyze the changes of gene expression triggered by 31-bp InDel during pollen development, two vectors (p385-In/Del::OsSUT3-GUS) were constructed and then stably introduced into rice. Histochemical and quantitative real-time PCR (qRT-PCR) analysis of transgenic plants showed that 31-bp deletion drastically reduced the expressions of downstream genes, including both OsSUT3 and GUS in rice panicle at booting stage, especially that of OsSUT3. The transcriptome profile of two types of panicles at booting stage revealed a total of 1028 differentially expressed genes (DEGs) between 31-bp In and 31-bp Del transgenic plants. Further analyses showed that 397 of these genes were significantly enriched for the ‘metabolic process’ and ‘binding’. Among them, nineteen genes had a strong relationship with starch and sucrose metabolism and were identified as candidate genes potentially associated with the starch accumulation in rice pollen, which that was also verified via qRT-PCR. In summary, 31-bp InDel plays a crucial role not only in the regulation of downstream genes but in the expression of sucrose-starch metabolizing genes in multiple biological pathways, and provides a different regulation mechanism for sucrose metabolism in pollen. Full article

(This article belongs to the Special Issue Rice Molecular Breeding and Genetics 2.0)

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15 pages, 9672 KiB

Open AccessArticle

Introducing MdTFL1 Promotes Heading Date and Produces Semi-Draft Phenotype in Rice

by Van Giap Do, Youngsuk Lee, Seonae Kim, Sangjin Yang, Juhyeon Park and Gyungran Do

Int. J. Mol. Sci. 2023, 24(12), 10365; https://doi.org/10.3390/ijms241210365 - 20 Jun 2023

Cited by 2 | Viewed by 1398

Abstract

Flowering time (in rice, termed the heading date), plant height, and grain number are crucial agronomic traits for rice productivity. The heading date is controlled via environmental factors (day length and temperature) and genetic factors (floral genes). TERMINAL FLOWER 1 (TFL1) encodes a [...] Read more.

Flowering time (in rice, termed the heading date), plant height, and grain number are crucial agronomic traits for rice productivity. The heading date is controlled via environmental factors (day length and temperature) and genetic factors (floral genes). TERMINAL FLOWER 1 (TFL1) encodes a protein that controls meristem identity and participates in regulating flowering. In this study, a transgenic approach was used to promote the heading date in rice. We isolated and cloned apple MdTFL1 for early flowering in rice. Transgenic rice plants with antisense MdTFL1 showed an early heading date compared with wild-type plants. A gene expression analysis suggested that introducing MdTFL1 upregulated multiple endogenous floral meristem identity genes, including the (early) heading date gene family FLOWERING LOCUS T and MADS-box transcription factors, thereby shortening vegetable development. Antisense MdTFL1 also produced a wide range of phenotypic changes, including a change in overall plant organelles that affected an array of traits, especially grain productivity. The transgenic rice exhibited a semi-draft phenotype, increased leaf inclination angle, restricted flag leaf length, reduced spikelet fertility, and fewer grains per panicle. MdTFL1 plays a central role in regulating flowering and in various physiological aspects. These findings emphasize the role of TFL1 in regulating flowering in shortened breeding and expanding its function to produce plants with semi-draft phenotypes. Full article

(This article belongs to the Special Issue Rice Molecular Breeding and Genetics 2.0)

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13 pages, 4005 KiB

Open AccessArticle

Achieving High Expression of Cry in Green Tissues and Negligible Expression in Endosperm Simultaneously via rbcS Gene Fusion Strategy in Rice

by Hao Chen, Yuqing Huang, Mengnan Ye, Ya Wang, Xiuying He and Jumin Tu

Int. J. Mol. Sci. 2023, 24(10), 9045; https://doi.org/10.3390/ijms24109045 - 20 May 2023

Viewed by 1051

Abstract

To allay excessive public concern about the safety of transgenic foods, and to optimize insect-resistant genes expression to delay the evolution of resistance in pests, we developed a promising strategy to fuse the GOI (gene of interest) with OsrbcS (rice small subunit of [...] Read more.

To allay excessive public concern about the safety of transgenic foods, and to optimize insect-resistant genes expression to delay the evolution of resistance in pests, we developed a promising strategy to fuse the GOI (gene of interest) with OsrbcS (rice small subunit of ribulose bisphosphate carboxylase/oxygenase) in transgenic rice, which acted as a carrier, driven by the OsrbcS native promoter to sequester its expression in green tissues. Using eYFP as a trial, we reported a high-level accumulation of eYFP in green tissue and almost none in the seed and root of the fused construct compared to the non-fused construct. After applying this fusion strategy in insect-resistant rice breeding, recombinant OsrbcS-Cry1Ab/Cry1Ac expressed rice plants conferred high resistance to leaffolders and striped stem borers, among which two single-copy lines possessed normal agronomic performance in the field. Specifically, Cry1Ab/Cry1Ac protein levels in single-copy construct transgenic lines ranged from 1.8 to 11.5 µg g⁻¹ in the leaf, higher than the Actin I promoter-driven control, T51-1, about 1.78 µg g⁻¹ in the leaf, but negligible (only 0.00012–0.00117 µg g⁻¹) in endosperm by ELISA analysis. Our study provided a novel approach to creating Cry1Ab/Cry1Ac-free endosperm rice with a high level of insect-resistant protein in green tissues through the simultaneous usage of the OsrbcS promoter and OsrbcS as a fusion partner. Full article

(This article belongs to the Special Issue Rice Molecular Breeding and Genetics 2.0)

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