Oryza sativa
Ontology highlight
ABSTRACT:
OTHER RELATED OMICS DATASETS IN: E-GEOD-12069
PROVIDER: PRJNA113319 | ENA |
REPOSITORIES: ENA
Publications
Microarray analyses reveal that plant mutagenesis may induce more transcriptomic changes than transgene insertion.
Batista Rita R Saibo Nelson N Lourenço Tiago T Oliveira Maria Margarida MM
Proceedings of the National Academy of Sciences of the United States of America 20080226 9
Controversy regarding genetically modified (GM) plants and their potential impact on human health contrasts with the tacit acceptance of other plants that were also modified, but not considered as GM products (e.g., varieties raised through conventional breeding such as mutagenesis). What is beyond the phenotype of these improved plants? Should mutagenized plants be treated differently from transgenics? We have evaluated the extent of transcriptome modification occurring during rice improvement ...[more]
Similar Datasets
Project description:Inserting transgenes into bacterial chromosomes is generally quite involved, requiring a selection for cells carrying the insertion, usually for drug-resistance, or multiple cumbersome manipulations, or both. Several approaches use phage lambda red recombination, which allows for the possibility of mutagenesis of the transgene during a PCR step.We present a simple, rapid and highly efficient method for transgene insertion into the chromosome of Escherichia coli, Salmonella or Shigella at a benign chromosomal site using the site-specific recombination machinery of the transposon Tn7. This method requires very few manipulations. The transgene is cloned into a temperature-sensitive delivery plasmid and transformed into bacterial cells. Growth at the permissive temperature with induction of the recombination machinery leads to transgene insertion, and subsequent growth at the nonpermissive temperature cures the delivery plasmid. Transgene insertion is highly site-specific, generating insertions solely at the Tn7 attachment site and so efficient that it is not necessary to select for the insertion.This method is more efficient and straightforward than other techniques for transgene insertion available for E. coli and related bacteria, making moving transgenes from plasmids to a chromosomal location a simple matter. The non-requirement for selection is particularly well suited for use in development of unmarked strains for environmental release, such as live-vector vaccine strains, and also for promoter-fusion studies, and experiments in which every bacterial cell must express a transgene construct.
| S-EPMC1475584 | BioStudies
Project description:The development of new plant varieties by genetic modification aims at improving their features or introducing new qualities. However, concerns about the unintended effects of transgenes and negative environmental impact of genetically modified plants are an obstacle for the use of these plants in crops. To analyze the impact of transgenesis on plant genomes, we analyze three cucumber transgenic lines with an introduced <i>thaumatin II</i> gene. After genomes sequencing, we analyzed the transgene insertion site and performed variant prediction. As a result, we obtained similar number of variants for all analyzed lines (average of 4307 polymorphisms), with high abundance in one region of chromosome 4. According to SnpEff analysis, the presence of genomic variants generally does not influence the genome functionality, as less than 2% of polymorphisms have high impact. Moreover, analysis indicates that these changes were more likely induced by in vitro culture than by the transgenesis itself. The insertion site analysis shows that the region of transgene integration could cause changes in gene expression, by gene disruption or loss of promoter region continuity.<h4>Supplementary information</h4>The online version contains supplementary material available at 10.1007/s12298-021-00990-8.
| S-EPMC8139995 | BioStudies
Project description:Flowering is an indication of the transition from vegetative growth to reproductive growth and has considerable effects on the life cycle of soya bean (Glycine max). In this study, we employed the CRISPR/Cas9 system to specifically induce targeted mutagenesis of GmFT2a, an integrator in the photoperiod flowering pathway in soya bean. The soya bean cultivar Jack was transformed with three sgRNA/Cas9 vectors targeting different sites of endogenous GmFT2a via Agrobacterium tumefaciens-mediated transformation. Site-directed mutations were observed at all targeted sites by DNA sequencing analysis. T1-generation soya bean plants homozygous for null alleles of GmFT2a frameshift mutated by a 1-bp insertion or short deletion exhibited late flowering under natural conditions (summer) in Beijing, China (N39°58', E116°20'). We also found that the targeted mutagenesis was stably heritable in the following T2 generation, and the homozygous GmFT2a mutants exhibited late flowering under both long-day and short-day conditions. We identified some 'transgene-clean' soya bean plants that were homozygous for null alleles of endogenous GmFT2a and without any transgenic element from the T1 and T2 generations. These 'transgene-clean' mutants of GmFT2a may provide materials for more in-depth research of GmFT2a functions and the molecular mechanism of photoperiod responses in soya bean. They will also contribute to soya bean breeding and regional introduction.
| S-EPMC5785355 | BioStudies
Project description:The molecular mechanism underlying dormancy release and the induction of flowering remains poorly understood in woody plants. Mu-legacy is a valuable blueberry mutant, in which a transgene insertion caused increased expression of a RESPONSE REGULATOR 2-like gene (VcRR2). Mu-legacy plants, compared with nontransgenic 'Legacy' plants, show dwarfing, promotion of flower bud formation, and can flower under nonchilling conditions. We conducted transcriptomic comparisons in leaves, chilled and nonchilled flowering buds, and late-pink buds, and analyzed a total of 41 metabolites of six groups of hormones in leaf tissues of both Mu-legacy and 'Legacy' plants. These analyses uncovered that increased VcRR2 expression promotes the expression of a homolog of Arabidopsis thaliana ENT-COPALYL DIPHOSPHATE SYNTHETASE 1 (VcGA1), which induces new homeostasis of hormones, including increased gibberellin 4 (GA4) levels in Mu-legacy leaves. Consequently, increased expression of VcRR2 and VcGA1, which function in cytokinin responses and gibberellin synthesis, respectively, initiated the reduction in plant height and the enhancement of flower bud formation of the Mu-legacy plants through interactions of multiple approaches. In nonchilled flower buds, 29 differentially expressed transcripts of 17 genes of five groups of hormones were identified in transcriptome comparisons between Mu-legacy and 'Legacy' plants, of which 22 were chilling responsive. Thus, these analyses suggest that increased expression of VcRR2 was collectively responsible for promoting flower bud formation in highbush blueberry under nonchilling conditions. We report here for the first time the importance of VcRR2 to induce a suite of downstream hormones that promote flowering in woody plants.
| S-EPMC6804727 | BioStudies
Project description:Processing of double-stranded RNA precursors into small RNAs is an essential regulator of gene expression in plant development and stress response. Small RNA processing requires the combined activity of a functionally diverse group of molecular components. However, in most of the plant species, there are insufficient mutant resources to functionally characterize each encoding gene. Here, mutations in loci encoding protein machinery involved in small RNA processing in soya bean and Medicago truncatula were generated using the CRISPR/Cas9 and TAL-effector nuclease (TALEN) mutagenesis platforms. An efficient CRISPR/Cas9 reagent was used to create a bi-allelic double mutant for the two soya bean paralogous Double-stranded RNA-binding2 (GmDrb2a and GmDrb2b) genes. These mutations, along with a CRISPR/Cas9-generated mutation of the M. truncatula Hua enhancer1 (MtHen1) gene, were determined to be germ-line transmissible. Furthermore, TALENs were used to generate a mutation within the soya bean Dicer-like2 gene. CRISPR/Cas9 mutagenesis of the soya bean Dicer-like3 gene and the GmHen1a gene was observed in the T0 generation, but these mutations failed to transmit to the T1 generation. The irregular transmission of induced mutations and the corresponding transgenes was investigated by whole-genome sequencing to reveal a spectrum of non-germ-line-targeted mutations and multiple transgene insertion events. Finally, a suite of combinatorial mutant plants were generated by combining the previously reported Gmdcl1a, Gmdcl1b and Gmdcl4b mutants with the Gmdrb2ab double mutant. Altogether, this study demonstrates the synergistic use of different genome engineering platforms to generate a collection of useful mutant plant lines for future study of small RNA processing in legume crops.
2018-01-01 | S-EPMC5978873 | BioStudies
Project description:BACKGROUND: Plants engineered for abiotic stress tolerance may soon be commercialized. The engineering of these plants typically involves the manipulation of complex multigene networks and may therefore have a greater potential to introduce pleiotropic effects than the simple monogenic traits that currently dominate the plant biotechnology market. While research on unintended effects in transgenic plant systems has been instrumental in demonstrating the substantial equivalence of many transgenic plant systems, it is essential that such analyses be extended to transgenic plants engineered for stress tolerance. Drought-tolerant Arabidopsis thaliana were engineered through overexpression of the transcription factor ABF3 in order to investigate unintended pleiotropic effects. In order to eliminate position effects, the Cre/lox recombination system was used to create control plant lines that contain identical T-DNA insertion sites but with the ABF3 transgene excised. This additionally allowed us to determine if Cre recombinase can cause unintended effects that impact the transcriptome. RESULTS: Microarray analysis of control plant lines that underwent Cre-mediated excision of the ABF3 transgene revealed only two genes that were differentially expressed in more than one plant line, suggesting that the impact of Cre recombinase on the transcriptome was minimal. In the absence of drought stress, overexpression of ABF3 had no effect on the transcriptome, but following drought stress, differences were observed in the gene expression patterns of plants overexpressing ABF3 relative to control plants. Examination of the functional distribution of the differentially expressed genes revealed strong similarity indicating that unintended pathways were not activated. CONCLUSIONS: The action of ABF3 is tightly controlled in Arabidopsis. In the absence of drought stress, ectopic activation of drought response pathways does not occur. In response to drought stress, overexpression of ABF3 results in a reprogramming of the drought response, which is characterized by changes in the timing or strength of expression of some drought response genes, without activating any unexpected gene networks. These results illustrate that important gene networks are highly regulated in Arabidopsis and that engineering stress tolerance may not necessarily cause extensive changes to the transcriptome.
| S-EPMC2837038 | BioStudies
Project description:Agrobacterium-mediated transformation has been widely used in producing transgenic plants, and was recently used to generate "transgene-clean" targeted genomic modifications coupled with the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas9) system. Although tremendous variation in morphological and agronomic traits, such as plant height, seed fertility, and grain size, was observed in transgenic plants, the underlying mechanisms are not yet well understood, and the types and frequency of genetic variation in transformed plants have not been fully disclosed. To reveal the genome-wide variation in transformed plants, we sequenced the genomes of five independent T0 rice plants using next-generation sequencing (NGS) techniques. Bioinformatics analyses followed by experimental validation revealed the following: (1) in addition to transfer-DNA (T-DNA) insertions, three transformed plants carried heritable plasmid backbone DNA of variable sizes (855-5216 bp) and in different configurations with the T-DNA insertions (linked or apart); (2) each transgenic plant contained an estimated 338-1774 independent genetic variations (single nucleotide variations (SNVs) or small insertion/deletions); and (3) 2-6 new Tos17 insertions were detected in each transformed plant, but no other transposable elements or bacterial genomic DNA.
| S-EPMC5172603 | BioStudies
Project description:Recent evidence supports a role for RNA as a common pathogenic agent in both the “polyglutamine” and “untranslated” dominant expanded repeat disorders. One feature of all repeat sequences currently associated with disease is their predicted ability to form a hairpin secondary structure at the RNA level. In order to investigate mechanisms by which hairpin forming repeat RNAs could induce neurodegeneration, we have looked for alterations in gene transcripts as hallmarks of the cellular response to toxic hairpin repeat RNAs. Three disease associated repeat sequences - CAG, CUG and AUUCU - were specifically expressed in the neurons of Drosophila and resultant common, early, transcriptional changes assessed by microarray analyses. Transcripts that encode several components of the Akt/Gsk3-β signalling pathway were altered as a consequence of expression of these repeat RNAs, indicating that this pathway is a component of the neuronal response to these pathogenic RNAs and may represent an important common therapeutic target in this class of diseases. The heads from newly eclosed Male Drosophila were used for RNA extraction and profiling on Affymetrix Drosophile Genome 2.0 microarrays. Twenty samples were analysed, representing control and experimental lines. Two independently derived control lines were used in each exeriment, totaling 4 samples. The tri-nucleotide repeats (CAG, CUG, CAA) were represented by three independently derived two transgene insertion lines each in one experiment and two independent four transgene insertion lines each in a second experiment, totaling 15 samples. A single four transgene insertion line was analysed for AUUCU. Candidates were selected from the pool of transcripts which showed a ‘present’ call in either all independent lines for a particular repeat sequence, or in all samples for the elav- GAL4/+ control in that experiment. Where possible, T-tests were performed on raw values to determine samples that showed a significant difference with a P-value < 0.05.
| E-GEOD-27178 | BioStudies
Project description:The insect resistant maize YieldGard MON810 was studied to assess the extent to which introduction of a transgene may putatively alter the expression of endogenous genes by comparison of various GM lines vs. their non-transgenic counterparts. To assess the extent to which introduction of a transgene may putatively alter the expression of endogenous genes, GM lines of the insect resistant maize YieldGard MON810 were compared with non-transgenic counterparts. For a more in-depth study, high-throughput deep sequencing together with microarrays were used to compare the transcriptomes of immature embryos of the MON810 variety DKC6575, with a cryIA(b) transgene, and its near-isogenic variety Tietar, grown under controlled environmental conditions. This technique also allows characterisation of the transgenic mRNAs produced. 3'UTR-anchored mRNA-seq produced 1,802,571 sequences from DKC6575 and 1,170,973 from Tietar, which mapped to 14,712 and 14,854 unigenes, respectively. Up to 32 reads from the transgenic embryos matched to the synthetic cry1A(b) sequence, similar to medium-abundant mRNAs. Gene expression analysis, using the R-bioconductor packages EdgeR and DEseq, revealed 140 differentially expressed genes mainly involved in carbohydrate metabolism, protein metabolism and chromatin organisation. Comparison of the expression of 30 selected genes in two additional MON810 and near-isogenic variety pairs showed that most of them were differentially expressed in the three pairs of varieties analysed. Analysis of functional annotation and the precise moment of expression of the differentially expressed genes and physiological data obtained suggest a slight but significant delay in seed and plant maturation of MON810 plants. However, these transcriptomic changes were not associated to undesirable changes in the global phenotype or plant behaviour. Moreover, while most expression changes in MON810 immature embryos were maintained in other transgenic varieties, some gene expression was found to be modulated by the genetic background in which the transgene was introduced through conventional breeding programs.
| S-EPMC4072715 | BioStudies
Project description:Induced pluripotent stem (iPS) cell technology holds vast promises for a cure to the hemoglobinopathies. Constructs and methods to safely insert therapeutic genes to correct the genetic defect need to be developed. Site-specific insertion is a very attractive method for gene therapy because the risks of insertional mutagenesis are eliminated provided that a "safe harbor" is identified, and because a single set of validated constructs can be used to correct a large variety of mutations simplifying eventual clinical use. We report here the correction of α-thalassemia major hydrops fetalis in transgene-free iPS cells using zinc finger-mediated insertion of a globin transgene in the AAVS1 site on human chromosome 19. Homozygous insertion of the best of the 4 constructs tested led to complete correction of globin chain imbalance in erythroid cells differentiated from the corrected iPS cells.
| S-EPMC3543982 | BioStudies