scholarly journals Trackable Multiplex Recombineering (TRMR) and Next-Generation Genome Design Technologies: Modifying Gene Expression inE. coliby Inserting Synthetic DNA Cassettes and Molecular Barcodes

2018 ◽  
pp. 15-31 ◽  
Author(s):  
Emily F. Freed ◽  
Gur Pines ◽  
Carrie A. Eckert ◽  
Ryan T. Gill
Keyword(s):  
Gene Expression ◽  
Next Generation ◽  
Synthetic Dna ◽  
Molecular Barcodes ◽  
Genome Design

scholarly journals Modulation of Human Mesenchymal Stem Cells by Electrical Stimulation Using an Enzymatic Biofuel Cell

Catalysts ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 62
Author(s):  
Won-Yong Jeon ◽  
Seyoung Mun ◽  
Wei Beng Ng ◽  
Keunsoo Kang ◽  
Kyudong Han ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Electrical Stimulation ◽  
Regenerative Medicine ◽  
Cell Morphology ◽  
Electrical Current ◽  
Specific Gene ◽  
Next Generation ◽  
The Impact

Enzymatic biofuel cells (EBFCs) have excellent potential as components in bioelectronic devices, especially as active biointerfaces to regulate stem cell behavior for regenerative medicine applications. However, it remains unclear to what extent EBFC-generated electrical stimulation can regulate the functional behavior of human adipose-derived mesenchymal stem cells (hAD-MSCs) at the morphological and gene expression levels. Herein, we investigated the effect of EBFC-generated electrical stimulation on hAD-MSC cell morphology and gene expression using next-generation RNA sequencing. We tested three different electrical currents, 127 ± 9, 248 ± 15, and 598 ± 75 nA/cm2, in mesenchymal stem cells. We performed transcriptome profiling to analyze the impact of EBFC-derived electrical current on gene expression using next generation sequencing (NGS). We also observed changes in cytoskeleton arrangement and analyzed gene expression that depends on the electrical stimulation. The electrical stimulation of EBFC changes cell morphology through cytoskeleton re-arrangement. In particular, the results of whole transcriptome NGS showed that specific gene clusters were up- or down-regulated depending on the magnitude of applied electrical current of EBFC. In conclusion, this study demonstrates that EBFC-generated electrical stimulation can influence the morphological and gene expression properties of stem cells; such capabilities can be useful for regenerative medicine applications such as bioelectronic devices.

scholarly journals Differential Gene Expression in the Siphonophore Nanomia bijuga (Cnidaria) Assessed with Multiple Next-Generation Sequencing Workflows

PLoS ONE ◽  
2011 ◽  
Vol 6 (7) ◽  
pp. e22953 ◽  
Author(s):  
Stefan Siebert ◽  
Mark D. Robinson ◽  
Sophia C. Tintori ◽  
Freya Goetz ◽  
Rebecca R. Helm ◽  
...  
Keyword(s):  
Gene Expression ◽  
Next Generation Sequencing ◽  
Differential Gene Expression ◽  
Next Generation ◽  
Differential Gene ◽  
Generation Sequencing

scholarly journals Prime-boost vaccination regimens with INO-4800 and INO-4802 augment and broaden immune responses against SARS-CoV-2 in nonhuman primates

2021 ◽  
Author(s):  
Jewell N Walters ◽  
Blake Schouest ◽  
Ami Patel ◽  
Emma L Reuschel ◽  
Katherine Schultheis ◽  
...  
Keyword(s):  
Dna Vaccine ◽  
Nonhuman Primates ◽  
First Generation ◽  
Rhesus Macaques ◽  
Natural Infection ◽  
Next Generation ◽  
Synthetic Dna ◽  
One Year ◽  
Global Population

The enhanced transmissibility and immune evasion associated with emerging SARS-CoV-2 variants demands the development of next-generation vaccines capable of inducing superior protection amid a shifting pandemic landscape. Since a portion of the global population harbors some level of immunity from vaccines based on the original Wuhan-Hu-1 SARS-CoV-2 sequence or natural infection, an important question going forward is whether this immunity can be boosted by next-generation vaccines that target emerging variants while simultaneously maintaining long-term protection against existing strains. Here, we evaluated the immunogenicity of INO-4800, our synthetic DNA vaccine candidate for COVID-19 currently in clinical evaluation, and INO-4802, a next-generation DNA vaccine designed to broadly target emerging SARS-CoV-2 variants, as booster vaccines in nonhuman primates. Rhesus macaques primed over one year prior with the first-generation INO-4800 vaccine were boosted with either INO-4800 or INO-4802 in homologous or heterologous prime-boost regimens. Both boosting schedules led to an expansion of antibody responses which were characterized by improved neutralizing and ACE2 blocking activity across wild-type SARS-CoV-2 as well as multiple variants of concern. These data illustrate the durability of immunity following vaccination with INO-4800 and additionally support the use of either INO-4800 or INO-4802 in prime-boost regimens.

scholarly journals Chronic Gamma Irradiation Changes Phenotype and Gene Expression Partially Transmitted to Next-Generation Tomato Seedlings

Agronomy ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 1638
Author(s):  
Seong-Min Kim ◽  
Yeong Deuk Jo ◽  
Jae-In Chun ◽  
Jin-Baek Kim ◽  
Jin-Ho Kang
Keyword(s):  
Gene Expression ◽  
Gamma Irradiation ◽  
Gamma Rays ◽  
Mutation Breeding ◽  
Fruit Shape ◽  
Leaf Length ◽  
Trichome Density ◽  
Next Generation ◽  
Seed Number ◽  
And Control

Compared to the studies on acute irradiation of seeds, fewer studies have reported on the chronic irradiation of seedlings, especially in fruit-bearing vegetables. We examined the effects of chronic gamma irradiation on tomato (Solanum lycopersicum ‘Micro-Tom’) seedlings exposed to gamma rays (50, 100, 150, and 200 Gy) for 4 weeks. As the total dose of gamma rays increased, leaf length, trichome density, and seed number were reduced in the irradiated seedlings (M1). Additionally, a change in fruit shape was observed. Chronic gamma irradiation reduced the expression of two trichome-related genes and affected the expression levels of 11 reactive oxygen species (ROS)-related genes. We examined the transmittance of these effects using M2 plants. The trichome density and fruit shape were similar between M2 and control plants; however, a reduction in leaf length and seed number was detected in M2 plants. Interestingly, changes in the expression of four ROS-related genes (ZAT10, Mn-SOD, POD3, and RBOH1) found in M1 were detected in M2 plants. Thus, the changes in phenotype and gene expression induced by chronic gamma irradiation were transmitted to the next generation. Additionally, we found novel mutants from M2 plants, suggesting that chronic gamma irradiation may be considered in tomato mutation breeding.

scholarly journals DNA Methylation Landscape of 16 Canine Somatic Tissues by Methylation-Sensitive Restriction Enzyme-Based Next Generation Sequencing

2021 ◽  
Author(s):  
Jumpei Yamazaki ◽  
Yuki Matsumoto ◽  
Jaroslav Jelinek ◽  
Teita Ishizaki ◽  
Shingo Maeda ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Next Generation Sequencing ◽  
Restriction Enzyme ◽  
Companion Animals ◽  
Next Generation ◽  
Cpg Sites ◽  
Somatic Tissues ◽  
Methylation Sensitive Restriction Enzyme ◽  
Generation Sequencing

Abstract Background: DNA methylation plays important functions in gene expression regulation that is involved in individual development and various diseases. DNA methylation has been well studied in human and model organisms, but only limited data exist in companion animals like dog. Results: Using methylation-sensitive restriction enzyme-based next generation sequencing (Canine DREAM), we obtained canine DNA methylation maps from 16 somatic tissues. In total, we evaluated 130,861 CpG sites. The majority of CpG sites were either highly methylated (>70%, 52.5%-64.6% of all CpG sites analyzed) or unmethylated (<30%, 22.5%-28.0% of all CpG sites analyzed) which are methylation patterns similar to other species. The overall methylation status of CpG sites across the 32 methylomes were remarkably similar. However, the tissue types were clearly defined by principle component analysis and hierarchical clustering analysis with DNA methylome. We found 6416 CpG sites located closely at promoter region of genes and inverse correlation between DNA methylation and gene expression of these genes. Conclusions: Our study provides basic dataset for DNA methylation profiles in dogs.

243 GENE EXPRESSION PROFILING BY NEXT-GENERATION SEQUENCING OF cDNA SAMPLES FROM INNER CELL MASS AND TROPHECTODERM OF A SINGLE-DAY 12 PORCINE EMBRYO

2010 ◽  
Vol 22 (1) ◽  
pp. 279
Author(s):  
S. C. Isom ◽  
R. S. Prather
Keyword(s):  
Gene Expression ◽  
Next Generation Sequencing ◽  
Gene Expression Profiling ◽  
Expression Profiling ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Next Generation ◽  
Sequencing Technologies ◽  
Cdna Fragments ◽  
Generation Sequencing

Traditional microarray approaches to gene expression profiling often require RNA or cDNA amplification when working with extremely small or valuable tissue samples.This process is generally viewed as being undesirable because there is potential for bias to be introduced during amplification. Very recently, the so-called next-generation sequencing technologies were adapted for use in global gene expression profiling. Herein we report our efforts to apply these sequencing technologies to assess relative transcript abundances in pre-implantation-stage porcine embryos, without additional nucleic acid amplification before sequencing. As a proof-of-principle experiment, we have isolated total RNA from the embryonic disc (inner cell mass; ICM) and a small piece of trophectoderm (TE) from a Day 12 in vivo-produced embryo, which were estimated to be composed of 500 to 1000 cells each. The RNA was reverse transcribed using oligo-dT priming followed by second-strand cDNA synthesis. The double-stranded cDNA was then randomly sheared by sonication, and 10 ng of double-stranded cDNA fragments was used for sample preparation before sequencing. Prepared cDNA fragments (at 7 picomolar concentrations) were submitted for sequencing using the Illumina/Solexa platform as recommended. The millions of short (36 bp) reads generated by Illumina sequencing for each sample were then aligned to the swine UniGene database from NCBI, allowing for zero or one mismatches. Relative transcript abundances between cell types were profiled by considering the read counts for a given UniGene member as a percentage of the total number of reads generated for each cell type. It was demonstrated that approximately 11 000 and 9000 UniGene members were represented by a normalized average of 5 or more short reads per lane (0.001% of the total) in the ICM and TE samples, respectively. As expected, pluripotency factors, chromatin remodeling components, and cell-cell communication molecules were overrepresented in the ICM sample as compared with the TE sample. Conversely, epithelial determinants, ion transporters, and components of the steroid biosynthesis pathways were more abundant in the TE sample than in the ICM sample. Relative abundances of representative transcripts in these samples were verified by quantitative RT-PCR. In conclusion, we demonstrate the utility of next-generation sequencing technologies for gene expression profiling using even minute tissue samples and show that such analyses are possible even in species without a sequenced genome.

scholarly journals Synthetic protein-binding DNA sponge as a tool to tune gene expression and mitigate protein toxicity

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Xinyi Wan ◽  
Filipe Pinto ◽  
Luyang Yu ◽  
Baojun Wang
Keyword(s):  
Gene Expression ◽  
Gene Regulation ◽  
Protein Binding ◽  
Gene Networks ◽  
Dynamic Range ◽  
Gene Expression Regulation ◽  
Single Layer ◽  
Regulation Mechanism ◽  
Gene Circuits ◽  
Synthetic Dna

AbstractVersatile tools for gene expression regulation are vital for engineering gene networks of increasing scales and complexity with bespoke responses. Here, we investigate and repurpose a ubiquitous, indirect gene regulation mechanism from nature, which uses decoy protein-binding DNA sites, named DNA sponge, to modulate target gene expression in Escherichia coli. We show that synthetic DNA sponges can be designed to reshape the response profiles of gene circuits, lending multifaceted tuning capacities including reducing basal leakage by >20-fold, increasing system output amplitude by >130-fold and dynamic range by >70-fold, and mitigating host growth inhibition by >20%. Further, multi-layer DNA sponges for decoying multiple regulatory proteins provide an additive tuning effect on the responses of layered circuits compared to single-layer sponges. Our work shows synthetic DNA sponges offer a simple yet generalizable route to systematically engineer the performance of synthetic gene circuits, expanding the current toolkit for gene regulation with broad potential applications.

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