Rapidly evolving genetic features for desert adaptations in Stipagrostis pennata

Abstract Background Stipagrostis pennata is distributed in the mobile and semi-mobile sand dunes which can adapt well to extreme environments such as drought and high temperature. It is a pioneer plant species with potential for stabilizing sand dunes and ecological restoration. It can settle on moving sand dunes earlier than other desert plants. It can effectively improve the stability of sand dunes and help more plants settle down and increase plant diversity. However, despite its important ecological value, the genetic resources available for this species are limited. Results We used single-molecule real-time sequencing technology to obtain the complete full-length transcriptome of Stipagrostis pennata, including 90,204 unigenes with an average length of 2624 bp. In addition, the 5436 transcription factors identified in these unigenes are rich in stress resistance genes, such as MYB-related, C3H, bHLH, GRAS and HSF, etc., which may play a role in adapting to desert drought and strong wind stress. Intron retention events are abundant alternative splicing events. Stipagrostis pennata has experienced stronger positive selection, accelerating the fixation of advantageous variants. Thirty-eight genes, such as CPP/TSO1-like gene, have evolved rapidly and may play a role in material transportation, flowering and seed formation. Conclusions The present study captures the complete full-length transcriptome of Stipagrostis pennata and reveals its rapid evolution. The desert adaptation in Stipagrostis pennata is reflected in the regulation of gene expression and the adaptability of gene function. Our findings provide a wealth of knowledge for the evolutionary adaptability of desert grass species.

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The complexity of alternative splicing and landscape of tissue-specific expression in lotus (Nelumbo nucifera) unveiled by Illumina- and single-molecule real-time-based RNA-sequencing

DNA Research ◽

10.1093/dnares/dsz010 ◽

2019 ◽

Vol 26 (4) ◽

pp. 301-311 ◽

Cited By ~ 11

Author(s):

Yue Zhang ◽

Tonny Maraga Nyong'A ◽

Tao Shi ◽

Pingfang Yang

Keyword(s):

Alternative Splicing ◽

Real Time ◽

Single Molecule ◽

Intron Retention ◽

Critical Role ◽

Full Length ◽

Specific Expression ◽

Splice Isoforms ◽

Tissue Specific ◽

Genomic Studies

Abstract Alternative splicing (AS) plays a critical role in regulating different physiological and developmental processes in eukaryotes, by dramatically increasing the diversity of the transcriptome and the proteome. However, the saturation and complexity of AS remain unclear in lotus due to its limitation of rare obtainment of full-length multiple-splice isoforms. In this study, we apply a hybrid assembly strategy by combining single-molecule real-time sequencing and Illumina RNA-seq to get a comprehensive insight into the lotus transcriptomic landscape. We identified 211,802 high-quality full-length non-chimeric reads, with 192,690 non-redundant isoforms, and updated the lotus reference gene model. Moreover, our analysis identified a total of 104,288 AS events from 16,543 genes, with alternative 3ʹ splice-site being the predominant model, following by intron retention. By exploring tissue datasets, 370 tissue-specific AS events were identified among 12 tissues. Both the tissue-specific genes and isoforms might play important roles in tissue or organ development, and are suitable for ‘ABCE’ model partly in floral tissues. A large number of AS events and isoform variants identified in our study enhance the understanding of transcriptional diversity in lotus, and provide valuable resource for further functional genomic studies.

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Full-length SMRT transcriptome sequencing and microsatellite characterization in Paulownia catalpifolia

Scientific Reports ◽

10.1038/s41598-021-87538-8 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Yanzhi Feng ◽

Yang Zhao ◽

Jiajia Zhang ◽

Baoping Wang ◽

Chaowei Yang ◽

...

Keyword(s):

Single Molecule ◽

Average Length ◽

Full Length ◽

Timber Species ◽

Sequencing Data ◽

Genetic Studies ◽

Ssr Loci ◽

Average Distribution ◽

Nucleotide Repeats ◽

Distribution Distance

AbstractPaulownia catalpifolia is an important, fast-growing timber species known for its high density, color and texture. However, few transcriptomic and genetic studies have been conducted in P. catalpifolia. In this study, single-molecule real-time sequencing technology was applied to obtain the full-length transcriptome of P. catalpifolia leaves treated with varying degrees of drought stress. The sequencing data were then used to search for microsatellites, or simple sequence repeats (SSRs). A total of 28.83 Gb data were generated, 25,969 high-quality (HQ) transcripts with an average length of 1624 bp were acquired after removing the redundant reads, and 25,602 HQ transcripts (98.59%) were annotated using public databases. Among the HQ transcripts, 16,722 intact coding sequences, 149 long non-coding RNAs and 179 alternative splicing events were predicted, respectively. A total of 7367 SSR loci were distributed throughout 6293 HQ transcripts, of which 763 complex SSRs and 6604 complete SSRs. The SSR appearance frequency was 28.37%, and the average distribution distance was 5.59 kb. Among the 6604 complete SSR loci, 1–3 nucleotide repeats were dominant, occupying 97.85% of the total SSR loci, of which mono-, di- and tri-nucleotide repeats were 44.68%, 33.86% and 19.31%, respectively. We detected 112 repeat motifs, of which A/T (42.64%), AG/CT (12.22%), GA/TC (9.63%), GAA/TTC (1.57%) and CCA/TGG (1.54%) were most common in mono-, di- and tri-nucleotide repeats, respectively. The length of the repeat SSR motifs was 10–88 bp, and 4997 (75.67%) were ≤ 20 bp. This study provides a novel full-length transcriptome reference for P. catalpifolia and will facilitate the identification of germplasm resources and breeding of new drought-resistant P. catalpifolia varieties.

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

10.1101/330977 ◽

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.

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PacBio single-molecule long-read sequencing shed new light on the complexity of the Carex breviculmis transcriptome

BMC Genomics ◽

10.1186/s12864-019-6163-6 ◽

2019 ◽

Vol 20 (1) ◽

Cited By ~ 2

Author(s):

Ke Teng ◽

Wenjun Teng ◽

Haifeng Wen ◽

Yuesen Yue ◽

Weier Guo ◽

...

Keyword(s):

Single Molecule ◽

Average Length ◽

Transcriptome Profiling ◽

Full Length ◽

Regulation Mechanism ◽

Forage Production ◽

Smrt Sequencing ◽

Reference Transcriptome ◽

Long Read ◽

Ecological Conservation

Abstract Background Carex L., a grass genus commonly known as sedges, is distributed worldwide and contributes constructively to turf management, forage production, and ecological conservation. The development of next-generation sequencing (NGS) technologies has considerably improved our understanding of transcriptome complexity of Carex L. and provided a valuable genetic reference. However, the current transcriptome is not satisfactory mainly because of the enormous difficulty in obtaining full-length transcripts. Results In this study, we employed PacBio single-molecule long-read sequencing (SMRT) technology for whole-transcriptome profiling in Carex breviculmis. We generated 60,353 high-confidence non-redundant transcripts with an average length of 2302-bp. A total of 3588 alternative splicing events, and 1273 long non-coding RNAs were identified. Furthermore, 40,347 complete coding sequences were predicted, providing an informative reference transcriptome. In addition, the transcriptional regulation mechanism of C. breviculmis in response to shade stress was further explored by mapping the NGS data to the reference transcriptome constructed by SMRT sequencing. Conclusions This study provided a full-length reference transcriptome of C. breviculmis using the SMRT sequencing method for the first time. The transcriptome atlas obtained will not only facilitate future functional genomics studies but also pave the way for further selective and genic engineering breeding projects for C. breviculmis.

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SMRT Sequencing of the Full-Length Transcriptome of the Coelomactra antiquata

Frontiers in Genetics ◽

10.3389/fgene.2021.741243 ◽

2021 ◽

Vol 12 ◽

Author(s):

Aiping Deng ◽

Jinpeng Li ◽

Zebin Yao ◽

Gyamfua Afriyie ◽

Ziyang Chen ◽

...

Keyword(s):

Molecular Biology ◽

Single Molecule ◽

Reference Genome ◽

Average Length ◽

Full Length ◽

Smrt Sequencing ◽

Sequencing Technology ◽

Medicinal Value ◽

Biology Research

Coelomactra antiquata is an important aquatic economic shellfish with high medicinal value. However, because C. antiquata has no reference genome, a lot of molecular biology research cannot be carried out, so the analysis of its transcripts is an important step to study the regulatory genes of various substances in C. antiquata. In the present study, we conducted the first full-length transcriptome analysis of C. antiquata by using PacBio single-molecule real-time (SMRT) sequencing technology. The results identified a total of 39,209 unigenes with an average length of 2,732 bp, 23,338 CDSs, 251 AS events, 9,881 lncRNAs, 20,106 SSRs, and 2,316 TFs. Subsequently, 59.22% (23,220) of the unigenes were successfully annotated, of which 23,164, 18,711, 15,840, 13,534, and 13,474 unigenes could be annotated using NR, Swiss-prot, KOG, GO, and KEGG databases, respectively. This study lays the foundation for the follow-up research of molecular biology and provides a reference for studying the more medicinal value of C. antiquata.

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Full-length transcriptome analysis of Spodoptera frugiperda larval brain reveals detoxification genes

PeerJ ◽

10.7717/peerj.12069 ◽

2021 ◽

Vol 9 ◽

pp. e12069

Author(s):

Lei Yang ◽

Binglin Xing ◽

Fen Li ◽

Li Kui Wang ◽

Linlin Yuan ◽

...

Keyword(s):

Single Molecule ◽

Spodoptera Frugiperda ◽

Average Length ◽

Rapid Development ◽

Fall Armyworm ◽

Transcriptome Profiling ◽

Resistance Mechanisms ◽

Full Length ◽

Agricultural Pests ◽

Larval Brain

Background Spodoptera frugiperda (J. E. Smith), commonly known as fall armyworm (FAW), is one of the most destructive agricultural pests in the world and has posed a great threat to crops. The improper use of insecticides has led to rapid development of resistance. However, the genetic data available for uncovering the insecticide resistance mechanisms are scarce. Methods In this study, we used PacBio single-molecule real-time (SMRT) sequencing aimed at revealing the full-length transcriptome profiling of the FAW larval brain to obtain detoxification genes. Results A total of 18,642 high-quality transcripts were obtained with an average length of 2,371 bp, and 11,230 of which were successfully annotated in six public databases. Among these, 5,692 alternative splicing events were identified.

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

10.1101/770040 ◽

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.

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SMRT sequencing analysis reveals the full-length transcripts and alternative splicing patterns in Ananas comosus var. bracteatus

PeerJ ◽

10.7717/peerj.7062 ◽

2019 ◽

Vol 7 ◽

pp. e7062 ◽

Cited By ~ 4

Author(s):

Jun Ma ◽

Yixuan Xiang ◽

Yingyuan Xiong ◽

Zhen Lin ◽

Yanbin Xue ◽

...

Keyword(s):

Alternative Splicing ◽

Single Molecule ◽

Intron Retention ◽

Chlorophyll Biosynthesis ◽

Full Length ◽

Ananas Comosus ◽

Cdna Libraries ◽

Sequencing Analysis ◽

Genomic Complexity ◽

Leaf Phenotype

Background Ananas comosus var. bracteatus is an herbaceous perennial monocot cultivated as an ornamental plant for its chimeric leaves. Because of its genomic complexity, and because no genomic information is available in the public GenBank database, the complete structure of the mRNA transcript is unclear and there are limited molecular mechanism studies for Ananas comosus var. bracteatus. Methods Three size fractionated full-length cDNA libraries (1–2 kb, 2–3 kb, and 3–6 kb) were constructed and subsequently sequenced in five single-molecule real-time (SMRT) cells (2 cells, 2 cells, and 1 cell, respectively). Results In total, 19,838 transcripts were identified for alternative splicing (AS) analysis. Among them, 19,185 (96.7%) transcripts were functionally annotated. A total of 9,921 genes were identified by mapping the non-redundant isoforms to the reference genome. A total of 10,649 AS events were identified, the majority of which were intron retention events. The alternatively spliced genes had functions in the basic metabolism processes of the plant such as carbon metabolism, amino acid biosynthesis, and glycolysis. Fourteen genes related to chlorophyll biosynthesis were identified as having AS events. The distribution of the splicing sites and the percentage of conventional and non-canonical AS sites of the genes categorized in pathways related to the albino leaf phenotype (ko00860, ko00195, ko00196, and ko00710) varied greatly. The present results showed that there were 8,316 genes carrying at least one poly (A) site, which generated 21,873 poly (A) sites. These findings indicated that the quality of the gene structure and functional information of the obtained genome was greatly improved, which may facilitate further genetic study of Ananas comosus var. bracteatus.

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Genome-Wide Analysis of Light-Regulated Alternative Splicing in Artemisia annua L.

Frontiers in Plant Science ◽

10.3389/fpls.2021.733505 ◽

2021 ◽

Vol 12 ◽

Author(s):

Tingyu Ma ◽

Han Gao ◽

Dong Zhang ◽

Wei Sun ◽

Qinggang Yin ◽

...

Keyword(s):

Alternative Splicing ◽

Single Molecule ◽

Artemisia Annua ◽

Intron Retention ◽

Genetic Regulation ◽

Red Light ◽

Light Regulation ◽

Full Length ◽

Common Mechanism ◽

Genome Wide

Artemisinin is currently the most effective ingredient in the treatment of malaria, which is thus of great significance to study the genetic regulation of Artemisia annua. Alternative splicing (AS) is a regulatory process that increases the complexity of transcriptome and proteome. The most common mechanism of alternative splicing (AS) in plant is intron retention (IR). However, little is known about whether the IR isoforms produced by light play roles in regulating biosynthetic pathways. In this work we would explore how the level of AS in A. annua responds to light regulation. We obtained a new dataset of AS by analyzing full-length transcripts using both Illumina- and single molecule real-time (SMRT)-based RNA-seq as well as analyzing AS on various tissues. A total of 5,854 IR isoforms were identified, with IR accounting for the highest proportion (48.48%), affirming that IR is the most common mechanism of AS. We found that the number of up-regulated IR isoforms (1534/1378, blue and red light, respectively) was more than twice that of down-regulated (636/682) after treatment of blue or red light. In the artemisinin biosynthetic pathway, 10 genes produced 16 differentially expressed IR isoforms. This work demonstrated that the differential expression of IR isoforms induced by light has the potential to regulate sesquiterpenoid biosynthesis. This study also provides high accuracy full-length transcripts, which can be a valuable genetic resource for further research of A. annua, including areas of development, breeding, and biosynthesis of active compounds.

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Full-Length Transcriptome of Thalassiosira weissflogii as a Reference Resource and Mining of Chitin-Related Genes

Marine Drugs ◽

10.3390/md19070392 ◽

2021 ◽

Vol 19 (7) ◽

pp. 392

Author(s):

Haomiao Cheng ◽

Chris Bowler ◽

Xiaohui Xing ◽

Vincent Bulone ◽

Zhanru Shao ◽

...

Keyword(s):

Cell Wall ◽

Full Length ◽

Biosynthetic Pathways ◽

Glycosidic Linkage ◽

Thalassiosira Weissflogii ◽

Ecological Value ◽

Pacbio Sequencing ◽

Non Coding Rnas ◽

Chitin And Chitosan ◽

Simple Sequence

β-Chitin produced by diatoms is expected to have significant economic and ecological value due to its structure, which consists of parallel chains of chitin, its properties and the high abundance of diatoms. Nevertheless, few studies have functionally characterised chitin-related genes in diatoms owing to the lack of omics-based information. In this study, we first compared the chitin content of three representative Thalassiosira species. Cell wall glycosidic linkage analysis and chitin/chitosan staining assays showed that Thalassiosira weissflogii was an appropriate candidate chitin producer. A full-length (FL) transcriptome of T. weissflogii was obtained via PacBio sequencing. In total, the FL transcriptome comprised 23,362 annotated unigenes, 710 long non-coding RNAs (lncRNAs), 363 transcription factors (TFs), 3113 alternative splicing (AS) events and 3295 simple sequence repeats (SSRs). More specifically, 234 genes related to chitin metabolism were identified and the complete biosynthetic pathways of chitin and chitosan were explored. The information presented here will facilitate T. weissflogii molecular research and the exploitation of β-chitin-derived high-value enzymes and products.

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