scholarly journals Full-length Transcriptome Analysis of Pecan (Carya illinoinensis) Kernels

2021 ◽  
Author(s):  
Chengcai Zhang ◽  
Huadong Ren ◽  
Xiaohua Yao ◽  
Kailiang Wang ◽  
Jun Chang
Keyword(s):  
Single Molecule ◽  
Molecular Mechanisms ◽  
Draft Genome ◽  
Alternative Polyadenylation ◽  
Full Length ◽  
Functional Annotations ◽  
Predominant Type ◽  
Long Read ◽  
Non Coding Rnas ◽  
Novel Isoforms

Abstract Pecan is rich in bioactive components such as fatty acids and flavonoids and is an important nut type worldwide. Therefore, the molecular mechanisms of phytochemical biosynthesis in pecan are a focus of research. Recently, a draft genome and several transcriptomes have been published. However, the full-length mRNA transcripts remain unclear, and the regulatory mechanisms behind the quality components biosynthesis and accumulation have not been fully investigated. In this study, single-molecule long read sequencing technology was used to obtain full-length transcripts of pecan kernels. In total, 37 504 isoforms of 16 702 genes were mapped to the reference genome. The numbers of known isoforms, new isoforms, and novel isoforms were 9013 (24.03%), 26 080 (69.54%), and 2411 (6.51%), respectively. Over 80% of the transcripts (30 751, 81.99%) had functional annotations. A total of 15 465 alternative splicing (AS) events and 65 761 alternative polyadenylation events were detected; wherein, the retained intron was the predominant type (5652, 36.55%) of AS. Furthermore, 1894 long non-coding RNAs and 1643 transcription factors were predicted using bioinformatics methods. Finally, the structural genes associated with fatty acid (FA) and flavonoid biosynthesis were characterized. A high frequency of AS accuracy (70.31%) was observed in FA synthesis-associated genes. The present study provides a full-length transcriptome dataset of pecan kernels, which will significantly enhance the understanding of the regulatory basis of phytochemical biosynthesis during pecan kernel maturation.

scholarly journals Single-Molecule Long-Read Sequencing Reveals the Diversity of Full-Length Transcripts in Leaves of Gnetum (Gnetales)

2019 ◽  
Vol 20 (24) ◽  
pp. 6350 ◽  
Author(s):  
Nan Deng ◽  
Chen Hou ◽  
Fengfeng Ma ◽  
Caixia Liu ◽  
Yuxin Tian
Keyword(s):  
Single Molecule ◽  
Developmental Stages ◽  
Alternative Polyadenylation ◽  
Full Length ◽  
Stomatal Development ◽  
Rna Seq ◽  
Leaf Transcriptome ◽  
Long Read ◽  
Non Coding Rnas ◽  
A Site

The limitations of RNA sequencing make it difficult to accurately predict alternative splicing (AS) and alternative polyadenylation (APA) events and long non-coding RNAs (lncRNAs), all of which reveal transcriptomic diversity and the complexity of gene regulation. Gnetum, a genus with ambiguous phylogenetic placement in seed plants, has a distinct stomatal structure and photosynthetic characteristics. In this study, a full-length transcriptome of Gnetum luofuense leaves at different developmental stages was sequenced with the latest PacBio Sequel platform. After correction by short reads generated by Illumina RNA-Seq, 80,496 full-length transcripts were obtained, of which 5269 reads were identified as isoforms of novel genes. Additionally, 1660 lncRNAs and 12,998 AS events were detected. In total, 5647 genes in the G. luofuense leaves had APA featured by at least one poly(A) site. Moreover, 67 and 30 genes from the bHLH gene family, which play an important role in stomatal development and photosynthesis, were identified from the G. luofuense genome and leaf transcripts, respectively. This leaf transcriptome supplements the reference genome of G. luofuense, and the AS events and lncRNAs detected provide valuable resources for future studies of investigating low photosynthetic capacity of Gnetum.

scholarly journals Analysis of Transcripts and splice isoforms in Red Clover (Trifolium pratense L.) by single-molecule long-read sequencing

2018 ◽  
Author(s):  
Yuehui Chao ◽  
Jianbo Yuan ◽  
Sifeng Li ◽  
Siqiao Jia ◽  
Liebao Han ◽  
...  
Keyword(s):  
Single Molecule ◽  
Trifolium Pratense ◽  
Intron Retention ◽  
Draft Genome ◽  
Red Clover ◽  
Full Length ◽  
Splice Isoforms ◽  
Single Molecule Level ◽  
Long Read ◽  
Novel Isoforms

AbstractRed clover (Trifolium pratense L.) is an important cool-season legume plant, which is the most widely planted forage legume after alfalfa. Although a draft genome sequence was published already, the sequences and completed structure of mRNA transcripts remain unclear, which limit further explore on red clover. In this study, the red clover transcriptome was sequenced using single-molecule long-read sequencing to identify full-length splice isoforms, and 29,730 novel isoforms from known genes and 2,194 novel isoforms from novel genes were identified. A total of 5,492 alternative splicing events was identified and the majority of alter spliced events in red clover was corrected as intron retention. In addition, of the 15,229 genes detected by SMRT, 8,719 including 1,86,517 transcripts have at least one poly(A) site. Furthermore, we identified 4,333 long non-coding RNAs and 3,762 fusion transcripts. Our results show the feasibility of deep sequencing full-length RNA from red clover transcriptome on a single-molecule level.

scholarly journals Single-nucleus full-length RNA profiling in plants incorporates isoform information to facilitate cell type identification

2020 ◽  
Author(s):  
Yanping Long ◽  
Zhijian Liu ◽  
Jinbu Jia ◽  
Weipeng Mo ◽  
Liang Fang ◽  
...  
Keyword(s):  
Single Cell ◽  
Single Molecule ◽  
Large Scale ◽  
Alternative Polyadenylation ◽  
Full Length ◽  
Cell Type ◽  
Arabidopsis Root ◽  
Rna Profiling ◽  
Long Read ◽  
Single Nucleus

AbstractThe broad application of large-scale single-cell RNA profiling in plants has been restricted by the prerequisite of protoplasting. We recently found that the Arabidopsis nucleus contains abundant polyadenylated mRNAs, many of which are incompletely spliced. To capture the isoform information, we combined 10x Genomics and Nanopore long-read sequencing to develop a protoplasting-free full-length single-nucleus RNA profiling method in plants. Our results demonstrated using Arabidopsis root that nuclear mRNAs faithfully retain cell identity information, and single-molecule full-length RNA sequencing could further improve cell type identification by revealing splicing status and alternative polyadenylation at single-cell level.

scholarly journals Combination of Single-Molecule Long-Read Sequencing and Illumina Sequencing Revealed the Mechanism of Anthocyanins Accumulation in an Ornamental Grass, Pennisetum Setaceum ‘Rubrum’

2021 ◽  
Author(s):  
Lingyun Liu ◽  
Ke Teng ◽  
Xifeng Fan ◽  
Hui Zhang ◽  
Chao Han ◽  
...  
Keyword(s):  
Single Molecule ◽  
Molecular Mechanisms ◽  
Color Change ◽  
Enrichment Analysis ◽  
Underlying Mechanism ◽  
Full Length ◽  
Anthocyanin Accumulation ◽  
Sequencing Technology ◽  
Long Read ◽  
Pennisetum Setaceum

Abstract Pennisetum setaceum ‘Rubrum’ is an ornamental herb with purple leaves, and it is widely used in the construction of landscaping. However, the current next generation sequencing (NGS) transcriptome information is not satisfactory mainly because of the enormous difficulty in obtaining full-length transcripts. What’s more, the molecular mechanisms of anthocyanin accumulation have not been thoroughly studied. In this study, we used PacBio full-length transcriptome sequencing combined with NGS sequencing technology to conduct transcriptome analysis on leaves showing different colors at different stages to clarify the molecular mechanism involved in the color change of P. setaceum ‘Rubrum’. A total of 280,413 full-length non-chimeric reads (FLNC) sequences were obtained based on single-molecule long-read sequencing technology. We obtained 140,633 high quality (HQ) transcripts and 2,683 low quality (LQ) transcripts and identified 5,352 alternative splicing (AS). In addition, a total of 93,066 ORFs, including 57,457 full open links and 2,910 lncRNA sequences were screened out. Furthermore, a total of 10,795 differentially expressed genes were identified. Gene ontology (GO) cluster and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis revealed the underlying mechanism of anthocyanin accumulation. In this study, to our best knowledge, we provided the full-length transcriptome information of P. setaceum ‘Rubrum’ for the first time. The underlying mechanism of anthocyanin accumulation in P. setaceum ‘Rubrum’ was further discussed based on the newly generated transcriptome data. The information will not only facilitate the gene function studies but also pave the way for future breeding projects of Pennisetum setaceum .

scholarly journals PacBio Single-Molecule Long-Read Sequencing Provides New Light on the Complexity of Full-Length Transcripts in Cattle

2021 ◽  
Vol 12 ◽  
Author(s):  
Tianpeng Chang ◽  
Bingxing An ◽  
Mang Liang ◽  
Xinghai Duan ◽  
Lili Du ◽  
...  
Keyword(s):  
Single Molecule ◽  
Bos Taurus ◽  
Human Life ◽  
Alternative Polyadenylation ◽  
Transcriptome Profiling ◽  
Full Length ◽  
Consensus Sequences ◽  
Long Read ◽  
Huge Impact ◽  
Whole Transcriptome

Cattle (Bos taurus) is one of the most widely distributed livestock species in the world, and provides us with high-quality milk and meat which have a huge impact on the quality of human life. Therefore, accurate and complete transcriptome and genome annotation are of great value to the research of cattle breeding. In this study, we used error-corrected PacBio single-molecule real-time (SMRT) data to perform whole-transcriptome profiling in cattle. Then, 22.5 Gb of subreads was generated, including 381,423 circular consensus sequences (CCSs), among which 276,295 full-length non-chimeric (FLNC) sequences were identified. After correction by Illumina short reads, we obtained 22,353 error-corrected isoforms. A total of 305 alternative splicing (AS) events and 3,795 alternative polyadenylation (APA) sites were detected by transcriptome structural analysis. Furthermore, we identified 457 novel genes, 120 putative transcription factors (TFs), and 569 novel long non-coding RNAs (lncRNAs). Taken together, this research improves our understanding and provides new insights into the complexity of full-length transcripts in cattle.

scholarly journals Reconstruction and functional annotation of Ascosphaera apis full-length transcriptome via PacBio single-molecule long-read sequencing

2019 ◽  
Author(s):  
Dafu Chen ◽  
Yu Du ◽  
Xiaoxue Fan ◽  
Zhiwei Zhu ◽  
Haibin Jiang ◽  
...  
Keyword(s):  
Single Molecule ◽  
Average Length ◽  
Full Length ◽  
Ascosphaera Apis ◽  
Protein Coding ◽  
Entomogenous Fungi ◽  
Consensus Sequences ◽  
Long Read ◽  
Non Coding Rnas ◽  
Lower Expression

AbstractAscosphaera apis is a widespread fungal pathogen of honeybee larvae that results in chalkbrood disease, leading to heavy losses for the beekeeping industry in China and many other countries. This work was aimed at generating a full-length transcriptome of A. apis using PacBio single-molecule real-time (SMRT) sequencing. Here, more than 23.97 Gb of clean reads was generated from long-read sequencing of A. apis mecylia, including 464,043 circular consensus sequences (CCS) and 394,142 full-length non-chimeric (FLNC) reads. In total, we identified 174,095 high-confidence transcripts covering 5141 known genes with an average length of 2728 bp. We also discovered 2405 genic loci and 11,623 isoforms that have not been annotated yet within the current reference genome. Additionally, 16,049, 10,682, 4520 and 7253 of the discovered transcripts have annotations in the Non-redundant protein (Nr), Clusters of Eukaryotic Orthologous Groups (KOG), Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases. Moreover, 1205 long non-coding RNAs (lncRNAs) were identified, which have less exons, shorter exon and intron lengths, shorter transcript lengths, lower GC percent, lower expression levels, and fewer alternative splicing (AS) evens, compared with protein-coding transcripts. A total of 253 members from 17 transcription factor (TF) families were identified from our transcript datasets. Finally, the expression of A. apis isoforms was validated using a molecular approach. Overall, this is the first report of a full-length transcriptome of entomogenous fungi including A. apis. Our data offer a comprehensive set of reference transcripts and hence contributes to improving the genome annotation and transcriptomic study of A. apis.

scholarly journals PacBio Single-Molecule Long-Read Sequencing Reveals Genes Tolerating Manganese Stress in Schima superba Saplings

2021 ◽  
Vol 12 ◽  
Author(s):  
Fiza Liaquat ◽  
Muhammad Farooq Hussain Munis ◽  
Samiah Arif ◽  
Urooj Haroon ◽  
Jianxin Shi ◽  
...  
Keyword(s):  
Single Molecule ◽  
Gene Annotation ◽  
Treated Group ◽  
Full Length ◽  
Open Reading Frames ◽  
Interacting Protein ◽  
Schima Superba ◽  
Long Read ◽  
First Time ◽  
Potential Tool

Schima superba (Theaceae) is a subtropical evergreen tree and is used widely for forest firebreaks and gardening. It is a plant that tolerates salt and typically accumulates elevated amounts of manganese in the leaves. With large ecological amplitude, this tree species grows quickly. Due to its substantial biomass, it has a great potential for soil remediation. To evaluate the thorough framework of the mRNA, we employed PacBio sequencing technology for the first time to generate S. Superba transcriptome. In this analysis, overall, 511,759 full length non-chimeric reads were acquired, and 163,834 high-quality full-length reads were obtained. Overall, 93,362 open reading frames were obtained, of which 78,255 were complete. In gene annotation analyses, the Kyoto Encyclopedia of Genes and Genomes (KEGG), Clusters of Orthologous Genes (COG), Gene Ontology (GO), and Non-Redundant (Nr) databases were allocated 91,082, 71,839, 38,914, and 38,376 transcripts, respectively. To identify long non-coding RNAs (lncRNAs), we utilized four computational methods associated with protein families (Pfam), Cooperative Data Classification (CPC), Coding Assessing Potential Tool (CPAT), and Coding Non-Coding Index (CNCI) databases and observed 8,551, 9,174, 20,720, and 18,669 lncRNAs, respectively. Moreover, nine genes were randomly selected for the expression analysis, which showed the highest expression of Gene 6 (Na_Ca_ex gene), and CAX (CAX-interacting protein 4) was higher in manganese (Mn)-treated group. This work provided significant number of full-length transcripts and refined the annotation of the reference genome, which will ease advanced genetic analyses of S. superba.

scholarly journals Allosteric activation of preformed EGF receptor dimers by binding of a single ligand

2021 ◽  
Author(s):  
Endang Purba ◽  
Ei-ichiro Saita ◽  
Reetesh Akhouri ◽  
Lars-Göran Öfverstedt ◽  
Gunnar Wilken ◽  
...  
Keyword(s):  
Ligand Binding ◽  
Single Molecule ◽  
Molecular Mechanisms ◽  
Egf Receptor ◽  
Electron Tomography ◽  
Growth Factor Receptor ◽  
Full Length ◽  
Cancer Therapies ◽  
Receptor Dimer

Abstract Aberrant activation of the epidermal growth factor receptor (EGFR) by mutations has been implicated in a variety of human cancers. Elucidation of the structure of the full-length receptor is essential to understand the molecular mechanisms underlying its activation. Unlike previously anticipated, here, we report that purified full-length EGFR adopts a homodimeric form in vitro before and after activation. Cryo-electron tomography analysis of the purified receptor also showed that the extracellular domains of the receptor dimer, which are conformationally flexible before activation, are stabilised by ligand binding. Consistently, optical single-molecule observation also demonstrated that binding of only one ligand activates the receptor dimer on the cell surface. Based on these results, we propose an allosteric model for the activation of EGFR dimers by ligand binding. Our results demonstrate how oncogenic mutations spontaneously activate the receptor and shed light on the development of novel cancer therapies.

scholarly journals A Full-Length Reference Floral Transcriptome of Boehmeria tricuspis Provides Insights into Apomeiosis and Polyploidy

2019 ◽  
Vol 2019 ◽  
pp. 1-13 ◽  
Author(s):  
Qing Tang ◽  
Ying Xu ◽  
Canhui Deng ◽  
Chaohua Cheng ◽  
Zhigang Dai ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Reproductive Mode ◽  
Full Length ◽  
Ploidy Levels ◽  
Reproductive Genes ◽  
Stress Response Genes ◽  
Long Read ◽  
Floral Transcriptome ◽  
Female Flowers ◽  
Significant Factors

Boehmeria tricuspis (Hance) Makino constitutes a hardy herbaceous or shrubby perennial native to East Asia that includes different ploidy levels and reproductive modes (diplosporous to sexual). Although several apomeiosis-associated genes have been described, the genetic control and molecular mechanisms underlying apomeiosis remain poorly understood. Moreover, the basis of the correlation between polyploidy and apomixis has not yet been clarified. We utilized long-read sequencing to produce a full-length reference floral transcriptome of B. tricuspis. Based on the generated database, gene expression of the female flowers of different ploidy levels and reproductive mode cytotypes was compared. Overall, 1,387 genes related to apomeiosis, 217 genes related to ploidy, and 9 genes associated with both apomixis and ploidy were identified. Gene Ontology analyses of this set of transcripts indicated reproductive genes, especially those related to “cell differentiation” and “cell cycle process,” as significant factors regulating apomeiosis. Furthermore, our results suggested that different expressions of stress response genes might be important in the preparation for apomeiosis transition. In addition, our observations indicated that the expression of apomeiosis may not depend on polyploidy but rather on deregulation of the sexual pathway in B. tricuspis.

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