scholarly journals Selective Enrichment of A-to-I Edited Transcripts from Cellular RNA Using Endonuclease V

2020 ◽  
Vol 142 (11) ◽  
pp. 5241-5251 ◽  
Author(s):  
Steve D. Knutson ◽  
Robert A. Arthur ◽  
H. Richard Johnston ◽  
Jennifer M. Heemstra
Keyword(s):  
Selective Enrichment ◽  
Endonuclease V

scholarly journals Selective enrichment of A-to-I edited transcripts from cellular RNA using Endonuclease V

2019 ◽  
Author(s):  
Steve D. Knutson ◽  
Jennifer M. Heemstra
Keyword(s):  
Escherichia Coli ◽  
Binding Affinity ◽  
Cost Effective ◽  
Effective Strategy ◽  
Robust Method ◽  
Rna Modifications ◽  
Selective Enrichment ◽  
Cost Effective Strategy ◽  
Endonuclease V

AbstractImmunoprecipitation enrichment has significantly improved the sensitivity and accuracy of detecting RNA modifications in the transcriptome. However, there are no existing methods for selectively isolating adenosine-to-inosine (A-to-I) edited RNAs. Here we show that Escherichia coli Endonuclease V (eEndoV), an inosine-cleaving enzyme, can be repurposed to bind and isolate A-to-I edited transcripts from cellular RNA through adjustment of cationic conditions. While Mg2+ is required for eEndoV catalysis, it has also been shown that similar levels of Ca2+ instead promote binding of inosine without cleavage. Leveraging these properties, we observe that Ca2+-supplemented eEndoV is highly specific for inosine in RNA and exhibits low nanomolar binding affinity. We then demonstrate EndoVIPER (Endonuclease Vinosine precipitation enrichment) as a facile and robust method to isolate A-to-I edited transcripts from cellular RNA. We envision the use of this approach as a straightforward and cost-effective strategy to enrich edited RNAs and detect A-to-I sites with improved sensitivity and fidelity.

Unravelling a microbial synergy to boost caproate production via carboxylates chain elongation with ethanol

2019 ◽  
Vol 26 (2) ◽  
pp. 63-71
Author(s):  
Ling Leng ◽  
Ying Wang ◽  
Peixian Yang ◽  
Takashi Narihiro ◽  
Masaru Konishi Nobu ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Volatile Fatty Acids ◽  
Microbial Consortia ◽  
Chain Elongation ◽  
Desulfovibrio Vulgaris ◽  
Rrna Gene ◽  
Selective Enrichment ◽  
Microbial Network ◽  
Cost Efficient ◽  
Rrna Gene Sequencing

Chain elongation of volatile fatty acids for medium chain fatty acids production (e.g. caproate) is an attractive approach to treat wastewater anaerobically and recover resource simultaneously. Undefined microbial consortia can be tailored to achieve chain elongation process with selective enrichment from anaerobic digestion sludge, which has advantages over pure culture approach for cost-efficient application. Whilst the metabolic pathway of the dominant caproate producer, Clostridium kluyveri, has been annotated, the role of other coexisting abundant microbiomes remained unclear. To this end, an ethanol-acetate fermentation inoculated with fresh digestion sludge at optimal conditions was conducted. Also, physiological study, thermodynamics and 16 S rRNA gene sequencing to elucidate the biological process by linking the system performance and dominant microbiomes were integrated. Results revealed a possible synergistic network in which C. kluyveri and three co-dominant species, Desulfovibrio vulgaris, Fusobacterium varium and Acetoanaerobium sticklandii coexisted. D. vulgaris and A. sticklandii (F. varium) were likely to boost the carboxylates chain elongation by stimulating ethanol oxidation and butyrate production through a syntrophic partnership with hydrogen (H2) serving as an electron messenger. This study unveils a synergistic microbial network to boost caproate production in mixed culture carboxylates chain elongation.

Fabrication of acylsemicarbazide-based porous organic polymer for selective enrichment of glycopeptides

2017 ◽  
Vol 35 (7) ◽  
pp. 688
Author(s):  
Hongwei WANG ◽  
Zhongshan LIU ◽  
Xiaojun PENG ◽  
Junjie OU ◽  
Mingliang YE
Keyword(s):  
Organic Polymer ◽  
Selective Enrichment ◽  
Porous Organic Polymer

Modelling of the micromorphology of the activated sludge floc: low do, low F/M bulking

1995 ◽  
Vol 31 (2) ◽  
pp. 235-243 ◽  
Author(s):  
Imre Takács ◽  
Ernö Fleit
Keyword(s):  
Activated Sludge ◽  
Bulk Concentration ◽  
Diffusion Limitation ◽  
Selective Enrichment ◽  
System A ◽  
Common Causes ◽  
Do Concentration ◽  
Soluble Substrate ◽  
Positive Feedback Effect ◽  
Do So

Two common causes of filamentous bulking of activated sludge are low dissolved oxygen (DO) concentration and low food to microorganism (F/M) ratio in the activated sludge system. A dynamic mathematical model was developed to simulate the population dynamics of two groups of bacteria, floc-formers and filaments within the microenvironment of the activated sludge floc. An arbitrary grid of 50 by 50 elements was applied to a hypothetical floc of maximum 100 μm in diameter. The concentration of DO and soluble substrate was calculated inside the floc core under different bulk concentration conditions in order to simulate the effect of heterogeneous, gradient-governed microenvironments on dual species composition. Dynamic simulation runs were performed to calculate the growth of the two morphological types of microorganisms inside the floc under diffusion governed conditions. The results indicate that the method accurately predicts the onset of excessive filamentous growth (directly linked to bulking) even when traditional models neglecting diffusion limitation fail to do so. The positive feedback effect of the non-random (unidirectional) growth on the selective enrichment of filamentous organisms under electron acceptor (DO) or soluble substrate (F/M) limited conditions is demonstrated.

Use of gene probes for the detection of quiescent enteric bacteria in marine and fresh waters

1995 ◽  
Vol 31 (5-6) ◽  
pp. 291-298
Author(s):  
Sally A. Anderson ◽  
Gillian D. Lewis ◽  
Michael N. Pearson
Keyword(s):  
Membrane Filtration ◽  
Enteric Bacteria ◽  
Probe Method ◽  
Detection Methods ◽  
23S Rrna ◽  
Specific Gene ◽  
Gene Probe ◽  
Selective Media ◽  
Selective Enrichment ◽  
E Coli

Specific gene probe detection methods that utilise a non-selective culturing step were tested for the ability to recognise the presence of quiescent enteric bacteria (Escherichia coli and Enterococcus faecalis ) within illuminated freshwater and seawater microcosms. An E. coli specific uidA gene probe and a 23S rRNA oligonucleotide probe for Enterococci were compared with recoveries using membrane filtration and incubation on selective media (mTEC and mE respectively). From these microcosm experiments a greater initial detection (from 4 hours to 1 day) of E. coli and Ent. faecalis using gene probe methods was observed. Additionally, a comparison of E. coli direct viable counts (DVC) in sunlight exposed microcosms with recoveries by selective media and gene probe methods revealed a large number of viable non-culturable cells. This suggests that enumeration of E. coli by a gene probe method is limited by the replication of the bacteria during the initial non-selective enrichment step. The detection of stressed Ent. faecalis by the oligonucleotide gene probe method was significantly greater than recovery on selective mE agar, indicating an Enterococci non-growth phase.

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