Transcriptomic analysis of floral initiation in litchi (Litchi chinensis Sonn.) based on de novo RNA sequencing

Hypoplastic Left Heart Syndrome (HLHS) is a complex multifactorial disease for which no definitive genetic causes have been found. Current genetic filtering strategies render lists of genes with unknown relevance in terms of pathogenesis. A complementary filter based on biological evidence would create a new approach to prioritize relevant candidate genes and mutations. In our study, 5 members of a nuclear family including a child with HLHS were evaluated using echocardiography and their genetic information was obtained through whole genome sequencing (WGS). Data filtering including rarity, functional impact and mode of inheritance was implemented, resulting in identification of 34 genes with recessive or de novo variants potentially involved in the pathogenesis of HLHS. Additionally, iPSC were derived from proband and parents and subjected to RNA-sequencing at the undifferentiated state and following spontaneous differentiation. Comparative transcriptional analyses identified genes differentially expressed in proband samples at each stage. These gene sets were used as an additional filter for the previously generated WGS data. This strategy revealed that out of 34 mutated genes originally identified, 10 displayed transcriptional differences in undifferentiated iPSC from the HLHS-affected individual while 16 out of 34 mutated genes showed significantly different expression levels in differentiated cells from proband. Furthermore, expression dynamics were studied during guided cardiac differentiation for the 9 genes fulfilling all applied criteria. Two genes not previously linked to HLHS, ELF4 and HSPG2 were found to behave significantly different in HLHS-iPSC when compared to control counterparts. In summary, filtering WGS data according to a new layer of transcriptional information that leverages iPSC plasticity allows prioritization of genes associated with HLHS in an in vitro model of disease.

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Comparative transcriptomic analysis of flowering induction in pitaya induced by supplementary lighting

10.21203/rs.2.9246/v1 ◽

2019 ◽

Author(s):

Rui Xiong ◽

Liu Chengli ◽

Min Xu ◽

Shuang-Shuang Wei ◽

Hua Tang

Keyword(s):

Time Course ◽

Molecular Mechanisms ◽

De Novo ◽

Floral Induction ◽

Transcriptomic Analysis ◽

Pcr Analysis ◽

Reverse Transcription Pcr ◽

Transcription Factor Genes ◽

Supplementary Lighting ◽

Transcriptomic Level

Abstract Background Pitayas are currently attracting considerable interest as a fruit with many health benefits. However, the lack of natural light after November in Hainan, China, severely restricts the production of pitaya in winter. To further explore the molecular mechanisms regulating flowering in pitaya, we used de novo RNA sequencing-based transcriptomic analysis for four stages of pitaya subjected to light induction. Results We assembled 68113 unigenes in total, comprising 29782 unigenes with functional annotations in the NR database, 20716 annotations in SwissProt, 18088 annotations in KOG, and 11059 annotations in KEGG. Comparison between different samples revealed different numbers of significantly differentially expressed genes (DEGs). A number of DEGs involved in energy metabolism-related processes and plant hormones were detected. Moreover, we discovered many CONSTANS-LIKE, FLOWERING LOCUS T and other DEGs involved in direct regulation of flowering, along with CDF and TCP, which function as typical transcription factor genes in the flowering process. At the transcriptomic level, we confirmed 13 DEGs with different functions in the time-course response to light-induced flowering by quantitative reverse-transcription PCR analysis. Conclusions These DEGs may include some key genes that control the floral-induction network, increasing our understanding of the molecular mechanism of floral regulation in pitaya. These findings will also aid the development of biotechnologies aimed at creating a variant of pitaya that is less sensitive to light conditions and blooms throughout the year.

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A general LC-MS-based RNA sequencing method for direct analysis of multiple-base modifications in RNA mixtures

Nucleic Acids Research ◽

10.1093/nar/gkz731 ◽

2019 ◽

Vol 47 (20) ◽

pp. e125-e125 ◽

Cited By ~ 8

Author(s):

Ning Zhang ◽

Shundi Shi ◽

Tony Z Jia ◽

Ashley Ziegler ◽

Barney Yoo ◽

...

Keyword(s):

Rna Sequencing ◽

De Novo ◽

Direct Analysis ◽

Complete Sequence ◽

Complete Understanding ◽

Sequencing Method ◽

Single Base Resolution ◽

Base Modifications ◽

Real Complex ◽

Usage Efficiency

Abstract A complete understanding of the structural and functional potential of RNA requires understanding of chemical modifications and non-canonical bases; this in turn requires advances in current sequencing methods to be able to sequence not only canonical ribonucleotides, but at the same time directly sequence these non-standard moieties. Here, we present the first direct and modification type-independent RNA sequencing method via introduction of a 2-dimensional hydrophobic end-labeling strategy into traditional mass spectrometry-based sequencing (2D HELS MS Seq) to allow de novo sequencing of RNA mixtures and enhance sample usage efficiency. Our method can directly read out the complete sequence, while identifying, locating, and quantifying base modifications accurately in both single and mixed RNA samples containing multiple different modifications at single-base resolution. Our method can also quantify stoichiometry/percentage of modified RNA versus its canonical counterpart RNA, simulating a real biological sample where modifications exist but may not be 100% at a particular site in the RNA. This method is a critical step towards fully sequencing real complex cellular RNA samples of any type and containing any modification type and can also be used in the quality control of modified therapeutic RNAs.

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Cytological and Gene Profile Expression Analysis Reveals Modification in Metabolic Pathways and Catalytic Activities Induce Resistance in Botrytis cinerea Against Iprodione Isolated From Tomato

International Journal of Molecular Sciences ◽

10.3390/ijms21144865 ◽

2020 ◽

Vol 21 (14) ◽

pp. 4865

Author(s):

Ambreen Maqsood ◽

Chaorong Wu ◽

Sunny Ahmar ◽

Haiyan Wu

Keyword(s):

Enzymatic Activity ◽

Rna Sequencing ◽

Aspartic Acid ◽

Mycelial Growth ◽

Plant Pathogens ◽

Fruits And Vegetables ◽

Transcriptomic Analysis ◽

Cell Membrane Permeability ◽

Wild Type ◽

Tomato Field

Grey mold is one of the most serious and catastrophic diseases, causing significant yield losses in fruits and vegetables worldwide. Iprodione is a broad spectrum agrochemical used as a foliar application as well as a seed protectant against many fungal and nematode diseases of fruits and vegetables from the last thirty years. The extensive use of agrochemicals produces resistance in plant pathogens and is the most devastating issue in food and agriculture. However, the molecular mechanism (whole transcriptomic analysis) of a resistant mutant of B. cinerea against iprodione is still unknown. In the present study, mycelial growth, sporulation, virulence, osmotic potential, cell membrane permeability, enzymatic activity, and whole transcriptomic analysis of UV (ultraviolet) mutagenic mutant and its wild type were performed to compare the fitness. The EC50 (half maximal effective concentration that inhibits the growth of mycelium) value of iprodione for 112 isolates of B. cinerea ranged from 0.07 to 0.87 µg/mL with an average (0.47 µg/mL) collected from tomato field of Guangxi Province China. Results also revealed that, among iprodione sensitive strains, only B67 strain induced two mutants, M0 and M1 after UV application. The EC50 of these induced mutants were 1025.74 μg/mL and 674.48 μg/mL, respectively, as compared to its wild type 1.12 μg/mL. Furthermore, mutant M0 showed higher mycelial growth sclerotia formation, virulence, and enzymatic activity than wild type W0 and M1 on potato dextrose agar (PDA) medium. The bctubA gene in the mutant M0 replaced TTC and GAT codon at position 593 and 599 by TTA and GAA, resulting in replacement of phenyl alanine into leucine (transversion C/A) and aspartic acid into glutamic acid (transversion T/C) respectively. In contrast, in bctubB gene, GAT codon at position 646 is replaced by AAT and aspartic acid converted into asparagine (transition G/A). RNA sequencing of the mutant and its wild type was performed without (M0, W0) and with iprodione treatment (M-ipro, W-ipro). The differential gene expression (DEG) identified 720 unigenes in mutant M-ipro than W-ipro after iprodione treatment (FDR ≤ 0.05 and log2FC ≥ 1). Seven DEGs were randomly selected for quantitative real time polymerase chain reaction to validate the RNA sequencing genes expression (log fold 2 value). The gene ontology (GO) enrichment and Kyoto encyclopedia genes and genomes (KEGG) pathway functional analyses indicated that DEG’s mainly associated with lysophopholipase, carbohydrate metabolism, amino acid metabolism, catalytic activity, multifunctional genes (MFO), glutathione-S transferase (GST), drug sensitivity, and cytochrome P450 related genes are upregulated in mutant type (M0, M-ipro) as compared to its wild type (W0, W-ipro), may be related to induce resistant in mutants of B. cinerea against iprodione.

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Molecular response of Sargassum vulgare to acidification at volcanic CO2 vents: insights from de novo transcriptomic analysis

Molecular Ecology ◽

10.1111/mec.14034 ◽

2017 ◽

Vol 26 (8) ◽

pp. 2276-2290 ◽

Cited By ~ 12

Author(s):

Amit Kumar ◽

Immacolata Castellano ◽

Francesco Paolo Patti ◽

Massimo Delledonne ◽

Hamada Abdelgawad ◽

...

Keyword(s):

De Novo ◽

Transcriptomic Analysis ◽

Molecular Response ◽

Sargassum Vulgare

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De novo RNA sequencing and analysis of the transcriptome of signalgrass (Urochloa decumbens) roots exposed to aluminum

Plant Growth Regulation ◽

10.1007/s10725-017-0291-2 ◽

2017 ◽

Vol 83 (1) ◽

pp. 157-170 ◽

Cited By ~ 7

Author(s):

Leonardo Rippel Salgado ◽

Rodolpho Lima ◽

Bruno Ferreira dos Santos ◽

Karina Tamie Shirakawa ◽

Mariane de Almeida Vilela ◽

...

Keyword(s):

Rna Sequencing ◽

De Novo ◽

Urochloa Decumbens

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A tissue level atlas of the healthy human virome

BMC Biology ◽

10.1186/s12915-020-00785-5 ◽

2020 ◽

Vol 18 (1) ◽

Cited By ~ 5

Author(s):

Ryuichi Kumata ◽

Jumpei Ito ◽

Kenta Takahashi ◽

Tadaki Suzuki ◽

Kei Sato

Keyword(s):

Gene Expression ◽

Virus Infection ◽

Rna Sequencing ◽

Human Gene ◽

Transcriptomic Analysis ◽

Immune Gene ◽

Healthy Individuals ◽

Human Gene Expression ◽

Somatic Tissues ◽

The Impact

Abstract Background Human-resident microbes can influence both health and disease. Investigating the microbiome using next-generation sequencing technology has revealed examples of mutualism and conflict between microbes and humans. Comparing to bacteria, the viral component of the microbiome (i.e., the “virome”) is understudied. Somatic tissues of healthy individuals are usually inaccessible for the virome sampling; therefore, there is limited understanding of the presence and distribution of viruses in tissues in healthy individuals and how virus infection associates with human gene expression and perturbs immunological homeostasis. Results To characterize the human virome in a tissue-specific manner, here we performed meta-transcriptomic analysis using the RNA-sequencing dataset from the Genotype-Tissue Expression (GTEx) Project. We analyzed the 8991 RNA-sequencing data obtained from 51 somatic tissues from 547 individuals and successfully detected 39 viral species in at least one tissue. We then investigated associations between virus infection and human gene expression and human disease onset. We detected some expected relationships; for instance, hepatitis C virus infection in the liver was strongly associated with interferon-stimulated gene upregulation and pathological findings of chronic hepatitis. The presence of herpes simplex virus type 1 in one subject’s brain strongly associated with immune gene expression. While torque teno virus was detected in a broad range of human tissues, it was not associated with interferon responses. Being notable in light of its association with lymphoproliferative disorders, Epstein-Barr virus infection in the spleen and blood was associated with an increase in plasma cells in healthy subjects. Human herpesvirus 7 was often detected in the stomach; intriguingly, it associated with the proportion of human leukocytes in the stomach as well as digestive gene expression. Moreover, virus infections in the local tissues associated with systemic immune responses in circulating blood. Conclusions To our knowledge, this study is the first comprehensive investigation of the human virome in a variety of tissues in healthy individuals through meta-transcriptomic analysis. Further investigation of the associations described here, and application of this analytical pipeline to additional datasets, will be useful to reveal the impact of viral infections on human health.

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