scholarly journals Recombinant Expression of Aldehyde Dehydrogenase 2 (ALDH2) inEscherichia coliNissle 1917 for Oral Delivery in ALDH2-Deficient Individuals

2019 ◽  
Author(s):  
Tim Ho ◽  
Catherine Chang ◽  
Justin Wu ◽  
Iris Huang ◽  
Leona Tsai ◽  
...  
Keyword(s):  
Aldehyde Dehydrogenase ◽  
Oral Delivery ◽  
Recombinant Expression ◽  
Data Availability ◽  
Supplementary Information ◽  
Aldehyde Dehydrogenase 2 ◽  
Design Data ◽  
E Coli ◽  
Competing Interests ◽  
Oral Conditions

AbstractTurning red after consuming alcohol may seem like a mere social inconvenience. Yet, this flushing response is caused by an accumulation of acetaldehyde, a carcinogenic intermediate of alcohol metabolism. Aldehyde dehydrogenase 2 (ALDH2) deficiency, the result of a point mutation, produces a less efficient ALDH2. The resulting accumulation of acetaldehyde greatly increases the risk of developing esophageal and head and neck cancers. In this study, we produced recombinant ALDH2 in the probioticE. coliNissle 1917, which successfully reduces acetaldehyde levels in simulated oral conditions. Packaged in a hard candy, the ALDH2-probiotic would remain in the mouth to specifically target salivary acetaldehyde. Using mathematical modeling, we also determined how much recombinant ALDH2 is needed to reduce elevated acetaldehyde levels.Financial DisclosureThis work was funded by Taipei American School. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Competing InterestsThe authors have declared that no competing interests exist.Ethics StatementN/AData AvailabilityYes – all data are fully available without restriction. Sequences for the plasmids used in this study are available through the Registry of Standard Biological Parts. Links to raw data are included in Supplementary Information.

scholarly journals Recombinant expression of Proteorhodopsin and biofilm regulators in Escherichia coli for nanoparticle binding and removal in a wastewater treatment model

2018 ◽  
Author(s):  
Justin Yang ◽  
Yvonne Wei ◽  
Catherine Yeh ◽  
Florence Liou ◽  
William Chen ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Wastewater Treatment ◽  
Recombinant Expression ◽  
High Surface ◽  
Volume Ratio ◽  
Data Availability ◽  
Supplementary Information ◽  
Design Data ◽  
Aquatic Life ◽  
Competing Interests

AbstractThe small size of nanoparticles is both an advantage and a problem. Their high surface-area-to-volume ratio enables novel medical, industrial, and commercial applications. However, their small size also allows them to evade conventional filtration during water treatment, posing health risks to humans, plants, and aquatic life. This project aims to remove nanoparticles during wastewater treatment using genetically modified Escherichia coli in two ways: 1) binding citrate-capped nanoparticles with the membrane protein Proteorhodopsin, and 2) trapping nanoparticles using Escherichia coli biofilm produced by overexpressing two regulators: OmpR234 and CsgD. We demonstrate experimentally that Escherichia coli expressing Proteorhodopsin binds to 60 nm citrate-capped silver nanoparticles. We also successfully upregulate biofilm production and show that Escherichia coli biofilms are able to trap 30 nm gold particles. Finally, both Proteorhodopsin and biofilm approaches are able to bind and remove nanoparticles in simulated wastewater treatment tanks. We envision integrating our trapping system in both rural and urban wastewater treatment plants to efficiently capture all nanoparticles before treated water is released into the environment.Financial DisclosureThis work was funded by the Taipei American School. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Competing InterestsThe authors have declared that no competing interests exist.Ethics StatementN/AData AvailabilityYes – all data are fully available without restriction. Sequences for the plasmids used in this study are available through the Registry of Standard Biological Parts. Links to raw data are included in Supplementary Information.

scholarly journals Quantification of DNA samples by Ethidium Bromide Spot Technique

2018 ◽  
Author(s):  
Jessica Zhao ◽  
Julian Zhao ◽  
Antoine Cummins ◽  
Tiffany Gonzalez ◽  
Grace Axler-DiPerte ◽  
...  
Keyword(s):  
Ethidium Bromide ◽  
Uv Light ◽  
Spot Size ◽  
Data Availability ◽  
Supplementary Information ◽  
Design Data ◽  
Dna Concentration ◽  
Standard Curve ◽  
Competing Interests ◽  
Concentration Method

AbstractAccurate and quick determination of DNA concentration is critical for the assembly of synthetic constructs, as well as a multitude of other experiments. We sought to optimize an under-utilized and inexpensive approach for determining DNA concentration: a spotting technique that uses the intercalating dye Ethidium Bromide. This technique does not require specialized equipment such as a spectrophotometer, but instead relies on visualization of dye-DNA complex fluorescence when excited by UV light. We modelled and tested a range of parameters for dye concentration and spot size, finding that 15uL spots with 1.0ug/mL Ethidium Bromide produced the most reliable standard curve. More importantly, we hope that our approach can help other labs optimize this protocol for their own experimental setup. Adoption of this technique may help enable development of iGEM teams in resource limited environments and laboratories which do not or cannot employ a satisfactory method for determining DNA concentration.Financial DisclosureWe obtained funding support from Canon Solutions America, and indirect support for this work through the CUNY Research Scholars Program. However, the funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Competing InterestsThe authors have declared that no competing interests exist.Ethics StatementN/AData AvailabilityYes – all data are fully available without restriction. The data can be found in the associated supplementary materials.Article TagsDNA, Ethidium Bromide, EtBr, Concentration, Method, UV, Gibson AssemblyHeader StatementThis work was assessed during the iGEM/PLOS Realtime Peer Review Jamboree on 23rd February 2018 and has been revised in response to the reviewers. Responses to Reviewers, and supplementary information (including Fig. S1) are available as separate files.

scholarly journals Characterization ofAzotobacter vinelandiiand Kits for Its Synthetic Biology Applications

2017 ◽  
Author(s):  
King Pong Leung ◽  
Jacky Fong Chuen Loo ◽  
Leo Chi U Seak ◽  
Tung Faat Lai ◽  
Kevin Yuk Lap Yip ◽  
...  
Keyword(s):  
Synthetic Biology ◽  
Azotobacter Vinelandii ◽  
Transmembrane Protein ◽  
Data Availability ◽  
Aerobic Bacterium ◽  
Design Data ◽  
Competing Interests ◽  
Enzymatic Cascades ◽  
Oxygen Sensitive ◽  
Anaerobic Environment

AbstractAzotobacter vinelandii, a Gram-negative aerobic bacterium with an intracellular anaerobic environment that maintains the oxygen-sensitive enzymatic cascades for nitrogen fixation, could be used to express oxygen-sensitive proteins. However, little is known about the properties ofA. vinelandiifor synthetic biology applications. We therefore first characterized and optimized the conditions for growing and screening BioBrick constructs inA. vinelandiiin the presence of 2 antibiotics, ampicillin and chloramphenicol, and then developed two sets of BioBricks for regulated protein expression. The first kit used T7 RNA polymerase, whose expression is under the control of a nitrogen-repressiblenifHpromoter. The commonly used T7-dependent system inEscherichia colican then be used inA. vinelandii. Because its intracellular anaerobic environment is favorable for processes such as magnetosome biogenesis, we attempted to migrate the biogenesis machineries from the magnetotactic bacteriumMagnetospirillum gryphiswaldensetoA. vinelandii. During this undertaking, another insertion kit construct was developed to allow protein conjugation onto magnetosomes. The kit consists ofmamC, a gene encoding a transmembrane protein on magnetosomes, and multiple restriction sites downstream ofmamCfor fusing a gene of interest. This insertion kit allows the attachment of any desired protein onto the magnetosome membrane by fusing with the mamC protein. We demonstrated the function of this kit by fusing mamC to a GFP nanobody. This kit will facilitate the conjugation of any target protein onto magnetosomes for downstream applications in the future.Financial DisclosureWe received sponsorship from the 2012–15 Teaching Development Grants Triennium, Faculty of Engineering and Biochemistry Program, School of Life Sciences, The Chinese University of Hong Kong. The funders had no role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript.Competing InterestsThe authors declare that no competing interests exist.Ethics StatementN/A.Data AvailabilityAll data are fully available without restriction.

Aldehyde Dehydrogenase 2 Polymorphism Is a Predictor of Smoking Cessation

2016 ◽  
pp. ntw316 ◽  
Author(s):  
Hiroyuki Masaoka ◽  
Silvano Gallus ◽  
Hidemi Ito ◽  
Miki Watanabe ◽  
Akira Yokomizo ◽  
...  
Keyword(s):  
Smoking Cessation ◽  
Aldehyde Dehydrogenase ◽  
Aldehyde Dehydrogenase 2

Long-term inhibition of ethanol intake by the administration of an aldehyde dehydrogenase-2 (ALDH2)-coding lentiviral vector into the ventral tegmental area of rats

2014 ◽  
Vol 20 (2) ◽  
pp. 336-344 ◽  
Author(s):  
Eduardo Karahanian ◽  
Mario Rivera-Meza ◽  
Lutske Tampier ◽  
María Elena Quintanilla ◽  
Mario Herrera-Marschitz ◽  
...  
Keyword(s):  
Ventral Tegmental Area ◽  
Aldehyde Dehydrogenase ◽  
Lentiviral Vector ◽  
Ethanol Intake ◽  
Aldehyde Dehydrogenase 2 ◽  
Ventral Tegmental
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