scholarly journals Full-Length Transcriptome Analysis of the ABCB, PIN/PIN-LIKES, and AUX/LAX Families Involved in Somatic Embryogenesis of Lilium pumilum DC. Fisch.

2020 ◽  
Vol 21 (2) ◽  
pp. 453 ◽  
Author(s):  
Shengli Song ◽  
Zhiping Wang ◽  
Yamin Ren ◽  
Hongmei Sun
Keyword(s):  
Somatic Embryogenesis ◽  
Single Molecule ◽  
Genetic Information ◽  
Auxin Transport ◽  
Regulatory Mechanism ◽  
Polar Auxin Transport ◽  
Regulatory Mechanisms ◽  
Smrt Sequencing ◽  
First Time ◽  
Somatic Embryo Induction

Plant cell totipotency is one of the 25 major topics in current scientific research, and somatic embryos are good experimental material for studying cell totipotency. Polar auxin transport plays an important regulatory role in somatic embryogenesis (SE). However, little is known about the auxin transport genes and their regulatory mechanisms in Lilium SE. In this study, we applied single-molecule real-time (SMRT) sequencing to Lilium pumilum DC. Fisch. for the first time and obtained a total of 119,649 transcripts, of which 14 encoded auxin transport genes. Correlation analyses between somatic embryo induction and gene expression under different treatments revealed that auxin transport genes, especially ATP-binding cassette (ABC) transporter B family member 21 (ABCB21) and PIN-FORMED (PIN) LIKES 7 (PILS7), may be key players in SE, and the necessary duration of picloram (PIC) treatment to induce SE is as short as 3 days. Our research provides valuable genetic information on Lilium pumilum, elucidating the candidate auxin transport genes involved in SE and their influencing factors. This study lays a foundation for elucidating the regulatory mechanism of auxin transport in SE.

Patterning during somatic embryogenesis in Scots pine in relation to polar auxin transport and programmed cell death

2011 ◽  
Vol 109 (3) ◽  
pp. 391-400 ◽  
Author(s):  
Malin Abrahamsson ◽  
Silvia Valladares ◽  
Emma Larsson ◽  
David Clapham ◽  
Sara von Arnold
Keyword(s):  
Cell Death ◽  
Somatic Embryogenesis ◽  
Programmed Cell Death ◽  
Scots Pine ◽  
Auxin Transport ◽  
Polar Auxin Transport

scholarly journals Understanding the diversity of DNA methylation in Mycobacterium tuberculosis

2020 ◽  
Author(s):  
Victor Ndhlovu ◽  
Anmol Kiran ◽  
Derek Sloan ◽  
Wilson Mandala ◽  
Marriot Nliwasa ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Mycobacterium Tuberculosis ◽  
Real Time ◽  
Single Molecule ◽  
Complete Loss ◽  
Phenotypic Heterogeneity ◽  
Epigenetic Variation ◽  
Smrt Sequencing ◽  
Underlying Mechanisms ◽  
First Time

AbstractAlthough Mycobacterium tuberculosis (Mtb) strains exhibit genomic homology of >99%, there is considerable variation in the phenotype. The underlying mechanisms of phenotypic heterogeneity in Mtb are not well understood but epigenetic variation is thought to contribute. At present the methylome of Mtb has not been completely characterized. We completed methylomes of 18 Mycobacterium tuberculosis (Mtb) clinical isolates from Malawi representing the largest number of Mtb genomes to be completed in a single study using Single Molecule Real Time (SMRT) sequencing to date. We replicate and confirm four methylation disrupting mutations in lineages of Mtb. For the first time we report complete loss of methylation courtesy of C758T (S253L) mutation in the MamB gene of Indo-oceanic lineage of Mtb. We also conducted a genomic and methylome comparison of the Malawian samples against a global sample. We confirm that methylation in Mtb is lineage specific although some unresolved issues still remain.

scholarly journals Single molecular real-time sequencing revealing the ovule abortion regulatory mechanisms in the female-sterile line of Pinus tabuliformis Carr.

2020 ◽  
Author(s):  
Zaixin Gong ◽  
Rui Han ◽  
Min Zhang ◽  
Qianquan Yang ◽  
Ming Zeng ◽  
...  
Keyword(s):  
Real Time ◽  
Single Molecule ◽  
Expression Patterns ◽  
Differentially Expressed ◽  
Regulatory Mechanisms ◽  
Cdna Libraries ◽  
Ovule Development ◽  
Smrt Sequencing ◽  
Ovule Abortion ◽  
Pinus Tabuliformis

Abstract Background Ovule abortion is a common phenomenon in plants, which has an essential significance in seed production. The development of the female gametophyte (FG) is one of the crucial processes of the life cycle, and FG dysplasia is a significant cause contribute to ovule abortion in many species. However, because it is difficult to acquire the mutant about ovule development in gymnosperms, previous studies of the ovule and FG development are mainly focused on angiosperms, especially in Arabidopsis thaliana. Thus the investigation on this field of gymnosperms remains unclear. Results In this study, we investigated the transcriptome data of wild-type (female fertile line, FER) and natural ovule abortion mutant (female sterile line, STE) of Pinus tabuliformis Carr. to evaluate the mechanism of ovule abortion during the process of free nuclear mitosis (FNM). Using single molecule real-time (SMRT) sequencing and next-generation sequencing (NGS), we obtained 202,869 (FER), 197,977 (STE) mapped full-length non-chimeric (FLNC) reads and analyzed 18 cDNA libraries. Based on the SMRT sequencing, we found that both GO annotation and KEGG pathway terms results were similar in FER and STE. NGS analysis further showed altogether 99 differentially expressed genes (DEGs) with opposite expression patterns during the FNM process in FER and STE. According to SMRT sequencing, we found that the number of isoforms and alternative splicing (AS) patterns are variable between FER and STE. Moreover, 5,530 and 5,096 potential simple sequence repeats (SSRs) were identified in FER and STE, respectively. Functional annotation results demonstrated that genes involved in energy metabolism, signal transduction, cell division, and stress response were differentially expressed in different lines, 9 pairs of which were confirmed by quantitative real-time PCR. Conclusion Taken together, these results provide some new insights about the ovule abortion in gymnosperms and further reveal the regulatory mechanisms of ovule development.

scholarly journals Characterization of DNA methylation in Malawian Mycobacterium tuberculosis clinical isolates

PeerJ ◽  
2020 ◽  
Vol 8 ◽  
pp. e10432
Author(s):  
Victor Ndhlovu ◽  
Anmol Kiran ◽  
Derek J. Sloan ◽  
Wilson Mandala ◽  
Marriott Nliwasa ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Single Molecule ◽  
Complete Loss ◽  
Clinical Isolates ◽  
Epigenetic Variation ◽  
Smrt Sequencing ◽  
Partial Loss ◽  
Underlying Mechanisms ◽  
First Time

Background Although Mycobacterium tuberculosis (Mtb) strains exhibit genomic homology of >99%, there is considerable variation in the phenotype. The underlying mechanisms of phenotypic heterogeneity in Mtb are not well understood but epigenetic variation is thought to contribute. At present the methylome of Mtb has not been completely characterized. Methods We completed methylomes of 18 Mycobacterium tuberculosis (Mtb) clinical isolates from Malawi representing the largest number of Mtb genomes to be completed in a single study using Single Molecule Real Time (SMRT) sequencing to date. Results We replicate and confirm four methylation disrupting mutations in 4 lineages of Mtb. For the first time we report complete loss of methylation courtesy of C758T (S253L) mutation in the MamB gene of Indo-oceanic lineage of Mtb. Additionally, we report a novel missense mutation G454A (G152S) in the MamA gene of the Euro-American lineage which could potentially be attributed to total disruption of methylation in the CCCAG motif but partial loss in a partner motif. Through a genomic and methylome comparative analysis with a global sample of sixteen, we report previously unknown mutations affecting the pks15/1 locus in L6 isolates. We confirm that methylation in Mtb is lineage specific although some unresolved issues still remain.

Somatic Embryogenesis and In vitro Plant Regeneration in Vigna trilobata (L.) Verd. - A Potential Pasture Legume

1970 ◽  
Vol 19 (1) ◽  
pp. 89-99
Author(s):  
K. Choudhary ◽  
M. Singh ◽  
M. S. Rathore ◽  
N. S. Shekhawat
Keyword(s):  
Somatic Embryogenesis ◽  
Growth Regulators ◽  
Somatic Embryos ◽  
Basal Medium ◽  
Long Term Study ◽  
Continuous Application ◽  
First Time ◽  
Pasture Legume

This long term study demonstrates for the first time that it is possible to propagate embryogenic Vigna trilobata and to subsequently initiate the differentiation of embryos into complete plantlets. Initiation of callus was possible on 2,4-D. Somatic embryos differentiated on modified MS basal nutrient medium with 1.0 mg/l  of 2,4-D and 0.5 mg/l  of Kn. Sustained cell division resulted in globular and heart shape stages of somatic embryos. Transfer of embryos on to a fresh modified MS basal medium with 0.5 mg/l of Kn and 0.5 mg/l of GA3 helped them to attain maturation and germination. However, the propagation of cells, as well as the differentiation of embryos, were inhibited by a continuous application of these growth regulators. For this reason, a long period on medium lacking these growth regulators was necessary before the differentiation of embryos occurred again. The consequences for improving the propagation of embryogenic cultures in Vigna species are discussed. Key words: Pasture  legume, Vigna trilobata, Globular, Heart shape, somatic embryogenesis D.O.I. 10.3329/ptcb.v19i1.4990 Plant Tissue Cult. & Biotech. 19(1): 89-99, 2009 (June)

scholarly journals Combined Transcriptome Analysis Reveals the Ovule Abortion Regulatory Mechanisms in the Female Sterile Line of Pinus tabuliformis Carr.

2021 ◽  
Vol 22 (6) ◽  
pp. 3138
Author(s):  
Zaixin Gong ◽  
Rui Han ◽  
Li Xu ◽  
Hailin Hu ◽  
Min Zhang ◽  
...  
Keyword(s):  
Real Time ◽  
Single Molecule ◽  
Regulatory Mechanisms ◽  
Cdna Libraries ◽  
Ovule Development ◽  
Ovule Abortion ◽  
Pinus Tabuliformis ◽  
Wild Type Female ◽  
Fertile Line ◽  
Next Generation Sequencing Ngs

Ovule abortion is a common phenomenon in plants that has an impact on seed production. Previous studies of ovule and female gametophyte (FG) development have mainly focused on angiosperms, especially in Arabidopsis thaliana. However, because it is difficult to acquire information about ovule development in gymnosperms, this remains unclear. Here, we investigated the transcriptomic data of natural ovule abortion mutants (female sterile line, STE) and the wild type (female fertile line, FER) of Pinus tabuliformis Carr. to evaluate the mechanism of ovule abortion during the process of free nuclear mitosis (FNM). Using single-molecule real-time (SMRT) sequencing and next-generation sequencing (NGS), 18 cDNA libraries via Illumina and two normalized libraries via PacBio, with a total of almost 400,000 reads, were obtained. Our analysis showed that the numbers of isoforms and alternative splicing (AS) patterns were significantly variable between FER and STE. The functional annotation results demonstrate that genes involved in the auxin response, energy metabolism, signal transduction, cell division, and stress response were differentially expressed in different lines. In particular, AUX/IAA, ARF2, SUS, and CYCB had significantly lower expression in STE, showing that auxin might be insufficient in STE, thus hindering nuclear division and influencing metabolism. Apoptosis in STE might also have affected the expression levels of these genes. To confirm the transcriptomic analysis results, nine pairs were confirmed by quantitative real-time PCR. Taken together, these results provide new insights into ovule abortion in gymnosperms and further reveal the regulatory mechanisms of ovule development.

Full-length-transcriptomic analysis in mice and human heart using Single-Molecule Real-time Sequencing (SMRT) identified 15 novel isoforms and a novel promoter region of PGC1-alpha

2020 ◽  
Vol 41 (Supplement_2) ◽  
Author(s):  
D Oehler ◽  
A Goedecke ◽  
A Spychala ◽  
K Lu ◽  
N Gerdes ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Single Molecule ◽  
High Fat Diet ◽  
Human Heart ◽  
Full Length ◽  
Funding Source ◽  
Smrt Sequencing ◽  
High Fat ◽  
Exon 1 ◽  
Novel Exon

Abstract Background Alternative splicing is a process by which exons within a pre-mRNA are joined or skipped, resulting in isoforms being encoded by a single gene. Alternative Splicing affecting transcription factors may have substantial impact on cellular dynamics. The PPARG Coactivator 1 Alpha (PGC1-α), is a major modulator in energy metabolism. Data from murine skeletal muscle revealed distinctive isoform patterns giving rise to different phenotypes, i.e. mitogenesis and hypertrophy. Here, we aimed to establish a complete dataset of isoforms in murine and human heart applying single-molecule real-time (SMRT)-sequencing as novel approach to identify transcripts without need for assembly, resulting in true full-length sequences. Moreover, we aimed to unravel functional relevance of the various isoforms during experimental ischemia reperfusion (I/R). Methods RNA-Isolation was performed in murine (C57Bl/6J) or human heart tissue (obtained during LVAD-surgery), followed by library preparation and SMRT-Sequencing. Bioinformatic analysis was done using a modified IsoSeq3-Pipeline and OS-tools. Identification of PGC1-α isoforms was fulfilled by similarity search against exonic sequences within the full-length, non-concatemere (FLNC) reads. Isoforms with Open-Reading-Frame (ORF) were manually curated and validated by PCR and Sanger-Sequencing. I/R was induced by ligature of the LAD for 45 min in mice on standard chow as well as on high-fat-high-sucrose diet. Area At Risk (AAR) and remote tissue were collected three and 16 days after I/R or sham-surgery (n=4 per time point). Promotor patterns were analyzed by qPCR. Results Deciphering the full-length transcriptome of murine and human heart resulted in ∼60000 Isoforms with 99% accuracy on mRNA-sequence. Focusing on murine PGC1-α-isoforms we discovered and verified 15 novel transcripts generated by hitherto unknown splicing events. Additionally, we identified a novel Exon 1 originating between the known promoters followed by a valid ORF, suggesting the discovery of a novel promoter. Remarkably, we found a homologous novel Exon1 in human heart, suggesting conservation of the postulated promoter. In I/R the AAR exhibited a significant lower expression of established and novel promoters compared to remote under standard chow 3d post I/R. 16d post I/R, the difference between AAR & Remote equalized in standard chow while remaining under High-Fat-Diet. Conclusion Applying SMRT-technique, we generated the first time a complete full-length-transcriptome of the murine and human heart, identifying 15 novel potentially coding transcripts of PGC1-α and a novel exon 1. These transcripts are differentially regulated in experimental I/R in AAR and remote myocardium, suggesting transcriptional regulation and alternative splicing modulating PGC1-α function in heart. Differences between standard chow and high fat diet suggest impact of impaired glucose metabolism on regulatory processes after myocardial infarction. Funding Acknowledgement Type of funding source: Public grant(s) – National budget only. Main funding source(s): Collaborative Research Centre 1116 (German Research Foundation)

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