scholarly journals A comparative transcriptional landscape of two castor cultivars obtained by single-molecule sequencing comparative analysis

2020 ◽  
Author(s):  
Wei Zhou ◽  
Yaxing Zhou ◽  
Guoli Zhu ◽  
Yun Wang ◽  
Zhibiao He ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Comparative Analysis ◽  
Alternative Splicing ◽  
Rna Sequencing ◽  
Transcriptome Sequencing ◽  
Full Length ◽  
Transcriptional Analysis ◽  
Sequencing Analysis ◽  
Short Read ◽  
Alternative Splicing Events

AbstractBackground and ObjectivesCastor (Ricinus communis L.) is an important non-edible oilseed crop. Lm type female strains and normal amphiprotic strains are important castor cultivars, and are mainly different in inflorescence structures and leaf shapes. To better understand the mechanisums underling these differences at the molecular level, we performed comparative transcriptional analysis.Materials and MethodsFull-length transcriptome sequencing and short-read RNA sequencing were employed.ResultsA total of 76,068 and 44,223 non-redundant transcripts were obtained from high-quality transcripts of Lm type female strains and normal amphiprotic strains, respectively. In Lm female strain and normal amphiprotic strains 51,613 and 20,152 alternative splicing events were found, respectively. There were 13,239 transcription factors identified from the full-length transcriptomes. Comparative analysis showed great different gene expression of common and unique transcription factors between the two cultivars. Meanwhile, functional analysis of isoform was conducted. Full-length sequences were used as a reference genome, and short-read RNA sequencing analysis was performed to conduct differential gene analysis. Furthermore, the function of DEGs were performed to annotation analysis.ConclusionsThe results revealed considerable difference and expression diversity between two cultivars, well beyond what was reported in previous studies, likely reflecting the differences in architecture between these two cultivars.HighlightUsing the full-length transcriptome sequencing technology, we performed comparative analysis of transcription factors of two castor cultivars, analyzed alternative splicing events, and identified their lncRNAs.

scholarly journals A Comparative Transcriptional Landscape of Two Castor Cultivars Obtained by Single-Molecule Sequencing Comparative Analysis

2021 ◽  
Vol 12 ◽  
Author(s):  
Yaxing Zhou ◽  
Guoli Zhu ◽  
Yun Wang ◽  
Zhibiao He ◽  
Wei Zhou
Keyword(s):  
Transcription Factors ◽  
Comparative Analysis ◽  
Rna Sequencing ◽  
Single Molecule ◽  
Ricinus Communis ◽  
Full Length ◽  
Transcriptional Analysis ◽  
Oilseed Crop ◽  
Sequencing Analysis ◽  
Short Read

Background and Objectives: Castor (Ricinus communis L.) is an important non-edible oilseed crop. Lm-type female strains and normal amphiprotic strains are important castor cultivars, and are mainly different in their inflorescence structures and leaf shapes. To better understand the mechanisms underlying these differences at the molecular level, we performed a comparative transcriptional analysis.Materials and Methods: Full-length transcriptome sequencing and short-read RNA sequencing were employed.Results: A total of 76,068 and 44,223 non-redundant transcripts were obtained from high-quality transcripts of Lm-type female strains and normal amphiprotic strains, respectively. In Lm-type female strains and normal amphiprotic strains, 51,613 and 20,152 alternative splicing events were found, respectively. There were 13,239 transcription factors identified from the full-length transcriptomes. Comparative analysis showed a great variety of gene expression of common and unique transcription factors between the two cultivars. Meanwhile, a functional analysis of the isoforms was conducted. The full-length sequences were used as a reference genome, and a short-read RNA sequencing analysis was performed to conduct differential gene analysis. Furthermore, the function of DEGs were performed to annotation analysis.Conclusion: The results revealed considerable differences and expression diversity between the two cultivars, well beyond what was reported in previous studies and likely reflecting the differences in architecture between these two cultivars.

scholarly journals The Full-Length Transcriptome of Spartina alterniflora Reveals the Complexity of High Salt Tolerance in Monocotyledonous Halophyte

2020 ◽  
Vol 61 (5) ◽  
pp. 882-896
Author(s):  
Wenbin Ye ◽  
Taotao Wang ◽  
Wei Wei ◽  
Shuaitong Lou ◽  
Faxiu Lan ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Salt Stress ◽  
Alternative Splicing ◽  
Salt Tolerance ◽  
Spartina Alterniflora ◽  
High Salt ◽  
Full Length ◽  
High Salt Tolerance ◽  
High Salt Stress ◽  
Alternative Splicing Events

Abstract Spartina alterniflora (Spartina) is the only halophyte in the salt marsh. However, the molecular basis of its high salt tolerance remains elusive. In this study, we used Pacific Biosciences (PacBio) full-length single-molecule long-read sequencing and RNA-seq to elucidate the transcriptome dynamics of high salt tolerance in Spartina by salt gradient experiments. High-quality unigenes, transcription factors, non-coding RNA and Spartina-specific transcripts were identified. Co-expression network analysis found that protein kinase-encoding genes (SaOST1, SaCIPK10 and SaLRRs) are hub genes in the salt tolerance regulatory network. High salt stress induced the expression of transcription factors but repressed the expression of long non-coding RNAs. The Spartina transcriptome is closer to rice than Arabidopsis, and a higher proportion of transporter and transcription factor-encoding transcripts have been found in Spartina. Transcriptome analysis showed that high salt stress induced the expression of carbohydrate metabolism, especially cell-wall biosynthesis-related genes in Spartina, and repressed its expression in rice. Compared with rice, high salt stress highly induced the expression of stress response, protein modification and redox-related gene expression and greatly inhibited translation in Spartina. High salt stress also induced alternative splicing in Spartina, while differentially expressed alternative splicing events associated with photosynthesis were overrepresented in Spartina but not in rice. Finally, we built the SAPacBio website for visualizing full-length transcriptome sequences, transcription factors, ncRNAs, salt-tolerant genes and alternative splicing events in Spartina. Overall, this study suggests that the salt tolerance mechanism in Spartina is different from rice in many aspects and is far more complex than expected.

scholarly journals Long-read RNA sequencing reveals widespread sex-specific alternative splicing in threespine stickleback fish

2020 ◽  
Author(s):  
Alice S. Naftaly ◽  
Shana Pau ◽  
Michael A. White
Keyword(s):  
Alternative Splicing ◽  
Rna Sequencing ◽  
Threespine Stickleback ◽  
Full Length ◽  
Transcription Start ◽  
Short Read ◽  
Transcription Start Sites ◽  
Alternative Transcripts ◽  
Gene Annotations ◽  
Long Read

AbstractAlternate isoforms contribute immensely to phenotypic diversity across eukaryotes. While short read RNA-sequencing has increased our understanding of isoform diversity, it is challenging to accurately detect full-length transcripts, preventing the identification of many alternate isoforms. Long-read sequencing technologies have made it possible to sequence full length alternative transcripts, accurately characterizing alternative splicing events, alternate transcription start and end sites, and differences in UTR regions. Here, we utilize PacBio long read RNA-sequencing (Iso-Seq) to examine the transcriptomes of five tissues in threespine stickleback fish (Gasterosteus aculeatus), a widely used genetic model species. The threespine stickleback fish has a refined genome assembly with gene annotations that are based on short-read RNA sequencing and predictions from coding sequence of other species. This suggests some of the existing annotations may be inaccurate or alternative transcripts may not be fully characterized. Using Iso-Seq we detected thousands of novel isoforms, indicating many isoforms are absent in the current Ensembl gene annotations. In addition, we refined many of the existing annotations within the genome. We noted many improperly positioned transcription start sites that were refined with long-read sequencing. The Iso-Seq predicted transcription start sites were more accurate, verified through ATAC-seq. We were also able to detect many alternative splicing events between sexes and across tissues. We found a substantial number of genes in both somatic and gonad tissue that had sex-specific isoforms. Our study highlights the power of long-read sequencing to study the complexity of transcriptomes, greatly improving genomic resources for the threespine stickleback fish.

scholarly journals Deep RNA sequencing reveals a high frequency of alternative splicing events in the fungus Trichoderma longibrachiatum

BMC Genomics ◽  
2015 ◽  
Vol 16 (1) ◽  
pp. 54 ◽  
Author(s):  
Bin-Bin Xie ◽  
Dan Li ◽  
Wei-Ling Shi ◽  
Qi-Long Qin ◽  
Xiao-Wei Wang ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Rna Sequencing ◽  
High Frequency ◽  
Trichoderma Longibrachiatum ◽  
Alternative Splicing Events

scholarly journals Analysis of Multiple Occurrences of Alternative Splicing Events in Arabidopsis thaliana Using Novel Sequenced Full-Length cDNAs

DNA Research ◽  
2009 ◽  
Vol 16 (3) ◽  
pp. 155-164 ◽  
Author(s):  
K. Iida ◽  
K. Fukami-Kobayashi ◽  
A. Toyoda ◽  
Y. Sakaki ◽  
M. Kobayashi ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Alternative Splicing ◽  
Full Length ◽  
Alternative Splicing Events

scholarly journals Long-read transcriptome sequencing analysis with IsoTools

2021 ◽  
Author(s):  
Matthias Lienhard ◽  
Twan van den Beucken ◽  
Florian Claiment ◽  
Stefan Boerno ◽  
Myriam Hochradel ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Transcriptome Sequencing ◽  
Sequencing Analysis ◽  
Differential Splicing ◽  
Accurate Analysis ◽  
Extensive Documentation ◽  
Long Read ◽  
Potential Applications ◽  
Transcriptional Changes ◽  
Analysis Package

Long-read transcriptome sequencing (LRTS) holds the promise to boost our understanding of alternative splicing. Recent advances in accuracy and throughput have diminished the major limitations and enabled the direct quantification of isoforms. Considering the complexity of the data and the broad range of potential applications, it is clear that highly flexible, accurate analysis tools are crucial. Here, we present IsoTools, a comprehensive Python-based analysis package, for the improvement of alternative and differential splicing analysis. IsoTools provides a comprehensive data structure that integrates genomic information from LRTS transcripts together with the reference annotation, and enables broad functionality to quality control, visualize and analyze the data. Additionally, we implemented a graph-based method for the identification of alternative splicing events and a statistical approach based on the beta binomial distribution for the detection of differential events. To demonstrate our methods, we generated PacBio Iso-Seq data of human hepatocytes treated with the HDAC inhibitor valproic acid, a compound known to induce widespread transcriptional changes. Contrasted with short read RNA-Seq of the same samples, this analysis shows that LRTS provides valuable additional insights for a better understanding of alternative splicing, in particular with respect to complex novel and differential splicing events. IsoTools is made available for the community along with extensive documentation at https://github.com/MatthiasLienhard/isotools.

scholarly journals Long-read transcriptome sequencing reveals abundant promoter diversity in distinct molecular subtypes of gastric cancer

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Kie Kyon Huang ◽  
Jiawen Huang ◽  
Jeanie Kar Leng Wu ◽  
Minghui Lee ◽  
Su Ting Tay ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Rna Sequencing ◽  
Molecular Subtypes ◽  
Full Length ◽  
Sequencing Data ◽  
Gastrointestinal Malignancies ◽  
Coding Sequences ◽  
Short Read ◽  
Long Read ◽  
Novel Isoforms

Abstract Background Deregulated gene expression is a hallmark of cancer; however, most studies to date have analyzed short-read RNA sequencing data with inherent limitations. Here, we combine PacBio long-read isoform sequencing (Iso-Seq) and Illumina paired-end short-read RNA sequencing to comprehensively survey the transcriptome of gastric cancer (GC), a leading cause of global cancer mortality. Results We performed full-length transcriptome analysis across 10 GC cell lines covering four major GC molecular subtypes (chromosomal unstable, Epstein-Barr positive, genome stable and microsatellite unstable). We identify 60,239 non-redundant full-length transcripts, of which > 66% are novel compared to current transcriptome databases. Novel isoforms are more likely to be cell line and subtype specific, expressed at lower levels with larger number of exons, with longer isoform/coding sequence lengths. Most novel isoforms utilize an alternate first exon, and compared to other alternative splicing categories, are expressed at higher levels and exhibit higher variability. Collectively, we observe alternate promoter usage in 25% of detected genes, with the majority (84.2%) of known/novel promoter pairs exhibiting potential changes in their coding sequences. Mapping these alternate promoters to TCGA GC samples, we identify several cancer-associated isoforms, including novel variants of oncogenes. Tumor-specific transcript isoforms tend to alter protein coding sequences to a larger extent than other isoforms. Analysis of outcome data suggests that novel isoforms may impart additional prognostic information. Conclusions Our results provide a rich resource of full-length transcriptome data for deeper studies of GC and other gastrointestinal malignancies.

scholarly journals Direct nanopore sequencing of mRNA reveals landscape of transcript isoforms in apicomplexan parasites

2020 ◽  
Author(s):  
V Vern Lee ◽  
Louise M. Judd ◽  
Aaron R. Jex ◽  
Kathryn E. Holt ◽  
Christopher J. Tonkin ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Rna Sequencing ◽  
Intron Retention ◽  
Transcript Level ◽  
Full Length ◽  
Sequencing Platform ◽  
Apicomplexan Parasites ◽  
Oxford Nanopore ◽  
Long Read ◽  
Widespread Phenomenon

AbstractAlternative splicing is a widespread phenomenon in metazoans by which single genes are able to produce multiple isoforms of the gene product. However, this has been poorly characterised in apicomplexans, a major phylum of some of the most important global parasites. Efforts have been hampered by atypical transcriptomic features, such as the high AT content of Plasmodium RNA, but also the limitations of short read sequencing in deciphering complex splicing events. In this study, we utilised the long read direct RNA sequencing platform developed by Oxford Nanopore Technologies (ONT) to survey the alternative splicing landscape of Toxoplasma gondii and Plasmodium falciparum. We find that while native RNA sequencing has a reduced throughput, it allows us to obtain full-length or near full-length transcripts with comparable quantification to Illumina sequencing. By comparing this data with available gene models, we find widespread alternative splicing, particular intron retention, in these parasites. Most of these transcripts contain premature stop codons, suggesting that in these parasites, alternative splicing represents a pathway to transcriptomic diversity, rather than expanding proteomic diversity. Moreover, alternative splicing rates are comparable between parasites, suggesting a shared splicing machinery, despite notable transcriptomic differences between the parasites. This work highlights a strategy in using long read sequencing to understand splicing events at the whole transcript level, and has implications in future interpretation of RNA-seq studies.

scholarly journals A gene toolbox for monitoring autophagy transcription

2021 ◽  
Vol 12 (11) ◽  
Author(s):  
Matteo Bordi ◽  
Rossella De Cegli ◽  
Beatrice Testa ◽  
Ralph A. Nixon ◽  
Andrea Ballabio ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Rna Sequencing ◽  
Real Time Pcr ◽  
Cellular Localization ◽  
Gene List ◽  
Sequencing Analysis ◽  
Functional Protein ◽  
Mtor Inhibition ◽  
Step Process ◽  
Autophagy Pathway

AbstractAutophagy is a highly dynamic and multi-step process, regulated by many functional protein units. Here, we have built up a comprehensive and up-to-date annotated gene list for the autophagy pathway, by combining previously published gene lists and the most recent publications in the field. We identified 604 genes and created main categories: MTOR and upstream pathways, autophagy core, autophagy transcription factors, mitophagy, docking and fusion, lysosome and lysosome-related genes. We then classified such genes in sub-groups, based on their functions or on their sub-cellular localization. Moreover, we have curated two shorter sub-lists to predict the extent of autophagy activation and/or lysosomal biogenesis; we next validated the “induction list” by Real-time PCR in cell lines during fasting or MTOR inhibition, identifying ATG14, ATG7, NBR1, ULK1, ULK2, and WDR45, as minimal transcriptional targets. We also demonstrated that our list of autophagy genes can be particularly useful during an effective RNA-sequencing analysis. Thus, we propose our lists as a useful toolbox for performing an informative and functionally-prognostic gene scan of autophagy steps.

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