Dataset Information

Transcript Complexity and New Insights of Restorer Line in CMS-D8 Cotton Through Full-Length Transcriptomic Analysis.

ABSTRACT: Hybrid utilization has proficiently increased crop production worldwide. The cytoplasmic male sterility (CMS) system has emerged as an efficient tool for commercial hybrid cotton seed production. The restorer line with dominant Rf₂ gene can restore the fertility of the CMS-D8 sterile line. However, the molecular mechanism of fertility restoration remains unclear in CMS-D8 cotton that limits wider utilization of three-line hybrid breeding. In our study, the Pacific Biosciences (PacBio) Iso-Seq technology was applied to understand fertility restoration mechanism of CMS-D8 cotton. In total, 228,106 full-length non-chimeric transcriptome sequences were obtained from anthers of developing flowering buds. The analysis results identified 3,174 novel isoforms, 2,597 novel gene loci, 652 long non-coding RNAs predicted from novel isoforms, 7,234 alternative splicing events, 114 fusion transcripts, and 1,667 genes with alternative polyadenylation. Specially, two novel genes associated with restoration function, Ghir_D05.742.1 and m64033_190821_201011/21103726/ccs were identified and showed significant higher levels of expression in restorer line than sterile and maintainer lines. Our comparative full-length transcriptome analysis provides new insights into the molecular function of Rf₂ fertility restorer gene. The results of this study offer a platform for fertility restoration candidate gene discovery in CMS-D8 cotton.

SUBMITTER: Feng J

PROVIDER: S-EPMC9253813 | biostudies-literature | 2022

REPOSITORIES: biostudies-literature

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Transcript Complexity and New Insights of Restorer Line in CMS-D8 Cotton Through Full-Length Transcriptomic Analysis.

Feng Juanjuan J Li Yongqi Y Zhang Jinfa J Zhang Meng M Zhang Xuexian X Shahzad Kashif K Guo Liping L Qi Tingxiang T Tang Huini H Wang Hailin H Qiao Xiuqin X Lin Zhongxu Z Xing Chaozhu C Wu Jianyong J

Frontiers in plant science 20220621

Hybrid utilization has proficiently increased crop production worldwide. The cytoplasmic male sterility (CMS) system has emerged as an efficient tool for commercial hybrid cotton seed production. The restorer line with dominant <i>Rf<sub>2</sub></i> gene can restore the fertility of the CMS-D8 sterile line. However, the molecular mechanism of fertility restoration remains unclear in CMS-D8 cotton that limits wider utilization of three-line hybrid breeding. In our study, the Pacific Biosciences ( ...[more]

PMID: 35800603

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Project description:BackgroundCytoplasmic male sterile (CMS) with cytoplasm from Gossypium Trilobum (D8) fails to produce functional pollen. It is useful for commercial hybrid cotton seed production. The restore line of CMS-D8 containing Rf2 gene can restore the fertility of the corresponding sterile line. This study combined the whole genome resequencing bulked segregant analysis (BSA) with high-throughput SNP genotyping to accelerate the physical mapping of Rf2 locus in CMS-D8 cotton.MethodsThe fertility of backcross population ((sterile line×restorer line)×maintainer line) comprising of 1623 individuals was investigated in the field. The fertile pool (100 plants with fertile phenotypes, F-pool) and the sterile pool (100 plants with sterile phenotypes, S-pool) were constructed for BSA resequencing. The selection of 24 single nucleotide polymorphisms (SNP) through high-throughput genotyping and the development insertion and deletion (InDel) markers were conducted to narrow down the candidate interval. The pentapeptide repeat (PPR) family genes and upregulated genes in restore line in the candidate interval were analysed by qRT-PCR.ResultsThe fertility investigation results showed that fertile and sterile separation ratio was consistent with 1:1. BSA resequencing technology, high-throughput SNP genotyping, and InDel markers were used to identify Rf2 locus on candidate interval of 1.48 Mb on chromosome D05. Furthermore, it was quantified in this experiment that InDel markers co-segregated with Rf2 enhanced the selection of the restorer line. The qRT-PCR analysis revealed PPR family gene Gh_D05G3391 located in candidate interval had significantly lower expression than sterile and maintainer lines. In addition, utilization of anther RNA-Seq data of CMS-D8 identified that the expression level of Gh_D05G3374 encoding NB-ARC domain-containing disease resistance protein in restorer lines was significantly higher than that in sterile and maintainer lines.ConclusionsThis study not only enabled us to precisely locate the restore gene Rf2 but also evaluated the utilization of InDel markers for marker assisted selection in the CMS-D8 Rf2 cotton breeding line. The results of this study provide an important foundation for further studies on the mapping and cloning of restorer genes.

Project description:BackgroundAs a vital type of noncoding RNAs, circular RNAs (circRNAs) play important roles in plant growth and development and stress response. However, little is known about the biological roles of circRNAs in regulating the stability of male fertility restoration for cytoplasmic male sterility (CMS) conditioned by Gossypium harknessii cytoplasm (CMS-D2) cotton under high-temperature (HT) stress.ResultsIn this study, RNA-sequencing and bioinformatics analysis were performed on pollen grains of isonuclear alloplasmic near-isogenic restorer lines NH [N(Rf1rf1)] and SH [S(Rf1rf1)] with obvious differences in fertility stability under HT stress at two environments. A total of 967 circRNAs were identified, with 250 differentially expressed under HT stress. We confirmed the back-splicing sites of eight selected circRNAs using divergent primers and Sanger sequencing. Tissue-specific expression patterns of five differentially expressed circRNAs (DECs) were also verified by RT-PCR and qRT-PCR. Functional enrichment and metabolic pathway analysis revealed that the parental genes of DECs were significantly enriched in fertility-related biological processes such as pollen tube guidance and cell wall organization, as well as the Pentose and glucuronate interconversions, Steroid biosynthesis, and N-Glycan biosynthesis pathways. Moreover, we also constructed a putative circRNA-mediated competing endogenous RNA (ceRNA) network consisting of 21 DECs, eight predicted circRNA-binding miRNAs, and their corresponding 22 mRNA targets, especially the two ceRNA modules circRNA346-miR159a-MYB33 and circRNA484-miR319e-MYB33, which might play important biological roles in regulating pollen fertility stability of cotton CMS-D2 restorer line under HT stress.ConclusionsThrough systematic analysis of the abundance, characteristics and expression patterns of circRNAs, as well as the potential functions of their parent genes, our findings suggested that circRNAs and their mediated ceRNA networks acted vital biological roles in cotton pollen development, and might be also essential regulators for fertility stability of CMS-D2 restorer line under heat stress. This study will open a new door for further unlocking complex regulatory mechanisms underpinning the fertility restoration stability for CMS-D2 in cotton.

Project description:Anther development and pollen fertility of cytoplasmic male sterility (CMS) conditioned by Gossypium harknessii cytoplasm (CMS-D2) restorer lines are susceptible to continuous high-temperature (HT) stress in summer, which seriously hinders the large-scale application of "three-line" hybrids in production. Here, integrated small RNA, transcriptome, degradome, and hormone profiling was performed to explore the roles of microRNAs (miRNAs) in regulating fertility stability in mature pollens of isonuclear alloplasmic near-isogenic restorer lines NH and SH under HT stress at two environments. A total of 211 known and 248 novel miRNAs were identified, of which 159 were differentially expressed miRNAs (DEMs). Additionally, 45 DEMs in 39 miRNA clusters (PmCs) were also identified, and most highly expressed miRNAs were significantly induced in SH under extreme HT, especially four MIR482 and six MIR6300 family miRNAs. PmC28 was located in the fine-mapped interval of the Rf1 gene and contained two DEMs, gra-miR482_L-2R + 2 and gma-miR2118a-3p_R + 1_1ss18TG. Transcriptome sequencing identified 6281 differentially expressed genes, of which heat shock protein (HSP)-related genes, such as HSP70, HSP22, HSP18.5-C, HSP18.2 and HSP17.3-B, presented significantly reduced expression levels in SH under HT stress. Through integrating multi-omics data, we constructed a comprehensive molecular network of miRNA-mRNA-gene-KEGG containing 35 pairs of miRNA/target genes involved in regulating the pollen development in response to HT, among which the mtr-miR167a_R + 1, tcc-miR167c and ghr-miR390a, tcc-miR396c_L-1 and ghr-MIR169b-p3_1ss6AG regulated the pollen fertility by influencing ARF8 responsible for the auxin signal transduction, ascorbate and aldarate metabolism, and the sugar and lipid metabolism and transport pathways, respectively. Further combination with hormone analysis revealed that HT-induced jasmonic acid signaling could activate the expression of downstream auxin synthesis-related genes and cause excessive auxin accumulation, followed by a cascade of auxin signal transduction, ultimately resulting in pollen abortion. The results provide a new understanding of how heat-responsive miRNAs regulate the stability of fertility restoration for CMS-D2 cotton under heat stress.

Project description:Adipose tissues have a central role in organisms, and adipose content is a crucial economic trait of poultry. Pekin duck is an ideal model to study the mechanism of abdominal and subcutaneous adipose deposition for its high ability of adipose synthesis and deposition. Alternative splicing contributes to functional diversity in abdominal and subcutaneous adipose. However, there has been no systematic analysis of the dynamics of differential alternative splicing of abdominal and subcutaneous adipose in Pekin duck. In our study, the Pacific Biosciences (PacBio) Iso-Seq technology was applied to explore the transcriptional complexity of abdominal and subcutaneous adipose in Pekin ducks. In total, 143,931 and 111,337 full-length non-chimeric transcriptome sequences of abdominal and subcutaneous adipocytes were obtained from 41.78 GB raw data, respectively. These data led us to identify 19,212 long non-coding RNAs (lncRNAs) and 74,571 alternative splicing events. In addition, combined with the next-generation sequencing technology, we correlated the structure and function annotation with the differential expression profiles of abdominal and subcutaneous adipose transcripts. This study identified lots of novel alternative splicing events and major transcripts of transcription factors related to adipose synthesis. STAT3 was reported as a vital gene for adipogenesis, and we found that its major transcript is STAT3-1, which may play a considerable role in the process of adipose synthesis in Pekin duck. This study greatly increases our understanding of the gene models, genome annotations, genome structures, and the complexity and diversity of abdominal and subcutaneous adipose in Pekin duck. These data provide insights into the regulation of alternative splicing events, which form an essential part of transcript diversity during adipogenesis in poultry. The results of this study provide an invaluable resource for studying alternative splicing and tissue-specific expression.

Dataset Information

Transcript Complexity and New Insights of Restorer Line in CMS-D8 Cotton Through Full-Length Transcriptomic Analysis.

Publications

Transcript Complexity and New Insights of Restorer Line in CMS-D8 Cotton Through Full-Length Transcriptomic Analysis.

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