CRAWview: for viewing splicing variation, gene families, and polymorphism in clusters of ESTs and full-length sequences

The plant CAtion/H+ eXchangers (CAX) proteins belong to Ca2+/cation antiporter (CaCA) superfamily. By using in silico methods, the CAX encoding genes in the genome of six legume species have been identified in this work. In examined legume genomes, the CAX genes belong to a small multigenic family. The number of the CAX genes in these legume species is 17 (soybean), 6 (common bean and C. cajan), 5 (M. truncatula and C. arietinum) and 3 genes (L. japonicus), respectively. The legume CAX genes vary in genomic full-length ranging from 1,213 to 11,561 base pairs. All of the genes exhibit introns (from 4 to 11 introns). Their deduced full-length protein sequences range from 248 to 718 amino acids. Theoretical pI values of most (39/42) of legume CAX proteins were less than 7. The secondary structure modelling of protein exhibit transmembrane helix region (from 3 to 11 regions). Half of all (23/42) included 11 transmembrane helix regions. Based on phylogeny analysis, all of the legume CAX were divided into two groups, A and B, each consisting of two subgroups. The phylogeny suggested an ancient gene duplication in the genome of legumes ancestry. The recent gene duplication even was only detected in the soybean genome after the speciation. The expression analysis showed that all of 3 L. japonicus CAX genes expressed in all examined tissues. However, the expression of C. cajan CAX genes was not detected. For each of 4 remaining legumes, the CAX genes were differed in their expression level depending on studied tissues. The tissue-specific expressions of some CAX genes were observed in 5 out of the 6 legume species, except C. cajan.

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Single molecule, full-length transcript sequencing provides insight into the extreme metabolism of ruby-throated hummingbird Archilochus colubris

10.1101/117218 ◽

2017 ◽

Cited By ~ 4

Author(s):

Rachael E. Workman ◽

Alexander M. Myrka ◽

Elizabeth Tseng ◽

G. William Wong ◽

Kenneth C. Welch ◽

...

Keyword(s):

Single Molecule ◽

De Novo ◽

Sequence Divergence ◽

Gene Families ◽

Full Length ◽

Sequencing Data ◽

Bioinformatics Pipeline ◽

Long Distance ◽

Migratory Flight ◽

Lack Of Information

AbstractHummingbirds can support their high metabolic rates exclusively by oxidizing ingested sugars, which is unsurprising given their sugar-rich nectar diet and use of energetically expensive hovering flight. However, they cannot rely on dietary sugars as a fuel during fasting periods, such as during the night, at first light, or when undertaking long-distance migratory flights, and must instead rely exclusively on onboard lipids. This metabolic flexibility is remarkable both in that the birds can switch between exclusive use of each fuel type within minutes and in that de novo lipogenesis from dietary sugar precursors is the principle way in which fat stores are built, sometimes at exceptionally high rates, such as during the few days prior to a migratory flight. The hummingbird hepatopancreas is the principle location of de novo lipogenesis and likely plays a key role in fuel selection, fuel switching, and glucose homeostasis. Yet understanding how this tissue, and the whole organism, achieves and moderates high rates of energy turnover is hampered by a fundamental lack of information regarding how genes coding for relevant enzymes differ in their sequence, expression, and regulation in these unique animals. To address this knowledge gap, we generated a de novo transcriptome of the hummingbird liver using PacBio full-length cDNA sequencing (Iso-Seq), yielding a total of 8.6Gb of sequencing data, or 2.6M reads from 4 different size fractions. We analyzed data using the SMRTAnalysis v3.1 Iso-Seq pipeline, including classification of reads and clustering of isoforms (ICE) followed by error-correction (Arrow). With COGENT, we clustered different isoforms into gene families to generate de novo gene contigs. We performed orthology analysis to identify closely related sequences between our transcriptome and other avian and human gene sets. We also aligned our transcriptome against the Calypte anna genome where possible. Finally, we closely examined homology of critical lipid metabolic genes between our transcriptome data and avian and human genomes. We confirmed high levels of sequence divergence within hummingbird lipogenic enzymes, suggesting a high probability of adaptive divergent function in the hepatic lipogenic pathways. Our results have leveraged cutting-edge technology and a novel bioinformatics pipeline to provide a compelling first direct look at the transcriptome of this incredible organism.

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Characterization and Analysis of the Full-Length Transcriptomes of Multiple Organs in Pseudotaxus chienii (W.C.Cheng) W.C.Cheng

International Journal of Molecular Sciences ◽

10.3390/ijms21124305 ◽

2020 ◽

Vol 21 (12) ◽

pp. 4305

Author(s):

Li Liu ◽

Zhen Wang ◽

Yingjuan Su ◽

Ting Wang

Keyword(s):

Expression Profiles ◽

Abiotic Factors ◽

Gene Families ◽

Full Length ◽

Conservation Strategies ◽

Soil Conditions ◽

Habitat Isolation ◽

Ecological Value ◽

Relict Species ◽

Expression Levels

Pseudotaxus chienii, a rare tertiary relict species with economic and ecological value, is a representative of the monotypic genus Pseudotaxus that is endemic to China. P. chienii can adapt well to habitat isolation and ecological heterogeneity under a variety of climate and soil conditions, and is able to survive in harsh environments. However, little is known about the molecular and genetic resources of this long-lived conifer. Herein, we sequenced the transcriptomes of four organs of P. chienii using the PacBio Isoform Sequencing and Illumina RNA Sequencing platforms. Based on the PacBio Iso-Seq data, we obtained 44,896, 58,082, 50,485, and 67,638 full-length unigenes from the root, stem, leaf, and strobilus, respectively, with a mean length of 2692 bp, and a mean N50 length of 3010.75 bp. We then comprehensively annotated these unigenes. The number of organ-specific expressed unigenes ranged from 4393 in leaf to 9124 in strobilus, suggesting their special roles in physiological processes, organ development, and adaptability in the different four organs. A total of 16,562 differentially expressed genes (DEGs) were identified among the four organs and clustered into six subclusters. The gene families related to biotic/abiotic factors, including the TPS, CYP450, and HSP families, were characterized. The expression levels of most DEGs in the phenylpropanoid biosynthesis pathway and plant–pathogen interactions were higher in the root than in the three other organs, suggesting that root constitutes the main organ of defensive compound synthesis and accumulation and has a stronger ability to respond to stress. The sequences were analyzed to predict transcription factors, long non-coding RNAs, and alternative splicing events. The expression levels of most DEGs of C2H2, C3H, bHLH, and bZIP families in the root and stem were higher than those in the leaf and strobilus, indicating that these TFs may play a crucial role in the survival of the root and stem. These results comprise the first comprehensive gene expression profiles obtained for different organs of P. chienii. Our findings will facilitate further studies on the functional genomics, adaptive evolution, and phylogeny of P. chienii, and lay the foundation for the development of conservation strategies for this endangered conifer.

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Full-Length Transcriptome Analysis of Four Different Tissues of Cephalotaxus oliveri

International Journal of Molecular Sciences ◽

10.3390/ijms22020787 ◽

2021 ◽

Vol 22 (2) ◽

pp. 787

Author(s):

Ziqing He ◽

Yingjuan Su ◽

Ting Wang

Keyword(s):

Single Molecule ◽

Global Climate ◽

Abiotic Factors ◽

Enrichment Analysis ◽

Gene Families ◽

Full Length ◽

Sequencing Data ◽

Smrt Sequencing ◽

Male Cone ◽

Temperature And Precipitation

Cephalotaxus oliveri is a tertiary relict conifer endemic to China, regarded as a national second-level protected plant in China. This species has experienced severe changes in temperature and precipitation in the past millions of years, adapting well to harsh environments. In view of global climate change and its endangered conditions, it is crucial to study how it responds to changes in temperature and precipitation for its conservation work. In this study, single-molecule real-time (SMRT) sequencing and Illumina RNA sequencing were combined to generate the complete transcriptome of C. oliveri. Using the RNA-seq data to correct the SMRT sequencing data, the four tissues obtained 63,831 (root), 58,108 (stem), 33,013 (leaf) and 62,436 (male cone) full-length unigenes, with a N50 length of 2523, 3480, 3181, and 3267 bp, respectively. Additionally, 35,887, 11,306, 36,422, and 25,439 SSRs were detected for the male cone, leaf, root, and stem, respectively. The number of long non-coding RNAs predicted from the root was the largest (11,113), and the other tissues were 3408 (stem), 3193 (leaf), and 3107 (male cone), respectively. Functional annotation and enrichment analysis of tissue-specific expressed genes revealed the special roles in response to environmental stress and adaptability in the different four tissues. We also characterized the gene families and pathways related to abiotic factors. This work provides a comprehensive transcriptome resource for C. oliveri, and this resource will facilitate further studies on the functional genomics and adaptive evolution of C. oliveri.

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The B-hordein prolamin family of barley

Genome ◽

10.1139/gen-2013-0016 ◽

2013 ◽

Vol 56 (3) ◽

pp. 179-185 ◽

Cited By ~ 9

Author(s):

Olin D. Anderson

Keyword(s):

Stop Codon ◽

Premature Stop Codon ◽

In Silico Analysis ◽

Gene Families ◽

Full Length ◽

C Terminus ◽

Est Assembly ◽

Silico Analysis ◽

Total Ests ◽

Relative Gene

The spectrum of B-hordein prolamins and genes in the single barley cultivar Barke is described from an in silico analysis of 1452 B-hordein ESTs and available genomic DNA. Eleven unique B-hordein proteins are derived from EST contigs. Ten contigs encode apparent full-length B-hordeins and the eleventh contains a premature stop codon that will lead to a truncated B-hordein. The 11 sequences are placed within the two previously described classes, i.e., the B1- and B3-type B-hordeins. The number of ESTs assigned to each sequence is used as an estimate of relative gene transcription and expression. Three of the sequences account for 79% of the total ESTs, with one sequence comprises 32% of the total ESTs and has a variant C-terminus caused by an undefined sequence change history near the 3′ coding terminus. The 70× difference in EST distribution among sequences points to the importance of understanding differential rates of expression within closely related gene families. Analysis of available genomic sequences confirms the EST assembly and reveals one full-length and two partial sequences of pseudogenes as evidenced by no matching ESTs for the sequences and premature stop codons and frame shifts.

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Reconstruction of the full-length transcriptome of cigar tobacco without a reference genome and characterization of anion channel/transporter transcripts

BMC Plant Biology ◽

10.1186/s12870-021-03091-6 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Hui Zhang ◽

Jingjing Jin ◽

Guoyun Xu ◽

Zefeng Li ◽

Niu Zhai ◽

...

Keyword(s):

Reference Genome ◽

Anion Channel ◽

Chloride Content ◽

Gene Families ◽

Full Length ◽

Raw Material ◽

Read Length ◽

Anion Channels ◽

Average Read Length ◽

Genes Encoding

Abstract Background Cigar wrapper leaves are the most important raw material of cigars. Studying the genomic information of cigar tobacco is conducive to improving cigar quality from the perspective of genetic breeding. However, no reference genome or full-length transcripts at the genome-wide scale have been reported for cigar tobacco. In particular, anion channels/transporters are of high interest for their potential application in regulating the chloride content of cigar tobacco growing on coastal lands, which usually results in relatively high Cl− accumulation, which is unfavorable. Here, the PacBio platform and NGS technology were combined to generate a full-length transcriptome of cigar tobacco used for cigar wrappers. Results High-quality RNA isolated from the roots, leaves and stems of cigar tobacco were subjected to both the PacBio platform and NGS. From PacBio, a total of 11,652,432 subreads (19-Gb) were generated, with an average read length of 1,608 bp. After corrections were performed in conjunction with the NGS reads, we ultimately identified 1,695,064 open reading frames including 21,486 full-length ORFs and 7,342 genes encoding transcription factors from 55 TF families, together with 2,230 genes encoding long non-coding RNAs. Members of gene families related to anion channels/transporters, including members of the SLAC and CLC families, were identified and characterized. Conclusions The full-length transcriptome of cigar tobacco was obtained, annotated, and analyzed, providing a valuable genetic resource for future studies in cigar tobacco.

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In Situ Localization of PPAR Gamma and Uncoupling Protein in Mouse Embryo Sections Using Digoxigenin-Labeled Riboprobes

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100162624 ◽

1996 ◽

Vol 54 ◽

pp. 32-33

Author(s):

C. Jennermann ◽

S. A. Kliewer ◽

D. C. Morris

Keyword(s):

Alkaline Phosphatase ◽

Room Temperature ◽

Uncoupling Protein ◽

Ppar Gamma ◽

Nuclear Hormone Receptor ◽

Full Length ◽

Northern Analysis ◽

Peroxisome Proliferator

Peroxisome proliferator-activated receptor gamma (PPARg) is a member of the nuclear hormone receptor superfamily and has been shown in vitro to regulate genes involved in lipid metabolism and adipocyte differentiation. By Northern analysis, we and other researchers have shown that expression of this receptor predominates in adipose tissue in adult mice, and appears first in whole-embryo mRNA at 13.5 days postconception. In situ hybridization was used to find out in which developing tissues PPARg is specifically expressed.Digoxigenin-labeled riboprobes were generated using the Genius™ 4 RNA Labeling Kit from Boehringer Mannheim. Full length PPAR gamma, obtained by PCR from mouse liver cDNA, was inserted into pBluescript SK and used as template for the transcription reaction. Probes of average size 200 base pairs were made by partial alkaline hydrolysis of the full length transcripts. The in situ hybridization assays were performed as described previously with some modifications. Frozen sections (10 μm thick) of day 18 mouse embryos were cut, fixed with 4% paraformaldehyde and acetylated with 0.25% acetic anhydride in 1.0M triethanolamine buffer. The sections were incubated for 2 hours at room temperature in pre-hybridization buffer, and were then hybridized with a probe concentration of 200μg per ml at 70° C, overnight in a humidified chamber. Following stringent washes in SSC buffers, the immunological detection steps were performed at room temperature. The alkaline phosphatase labeled, anti-digoxigenin antibody and detection buffers were purchased from Boehringer Mannheim. The sections were treated with a blocking buffer for one hour and incubated with antibody solution at a 1:5000 dilution for 2 hours, both at room temperature. Colored precipitate was formed by exposure to the alkaline phosphatase substrate nitrobluetetrazoliumchloride/ bromo-chloroindlylphosphate.

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