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Do-it-yourself CRISPR genome editing kits bring genetic engineering to your kitchen bench

A synthetic biologist from NASA plans to make CRISPR-based genetic engineering as accessible as a home science kit, so you can bio-hack yeast and bacteria on your kitchen bench.​...
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Single-step Precision Genome Editing in Yeast Using CRISPR-Cas9

Bio Protoc. 2018 Mar 20;8(6). pii: e2765. doi: 10.21769/BioProtoc.2765.
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Bio-Algorithmic Workflows for Standardized Synthetic Biology Constructs

Methods Mol Biol. 2018;1772:363-372. doi: 10.1007/978-1-4939-7795-6_20.
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CRISPR-Cas9-Mediated Genome Editing and Transcriptional Control in Yarrowia lipolytica

Methods Mol Biol. 2018;1772:327-345. doi: 10.1007/978-1-4939-7795-6_18.
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Tools for Biohackers: Here Come 3 Mini-Labs

Tools for Biohackers: Here Come 3 Mini-Labs | SynBioFromLeukipposInstitute | Scoop.it
These desktop gadgets should make DIY genetic engineering much easier...
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CRISPR Cas9 Nuclease Recruitment for Targeted Mutagenesis using Oligo SgRNA in Soybean (Glycine Max L) | SciTechnol

CRISPR Cas9 Nuclease Recruitment for Targeted Mutagenesis using Oligo SgRNA in Soybean (Glycine Max L) | SciTechnol | SynBioFromLeukipposInstitute | Scoop.it
The Modern era of genome engineering via targeted mutagenesis is now bringing further innovations in the field of Plant biology and biomedicines. RNA-guided endonucleases..
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'CaRROT' system uses light to turn genes on or off

'CaRROT' system uses light to turn genes on or off | SynBioFromLeukipposInstitute | Scoop.it
"It is possible that one day, by just exposing the tissues to light, we can heal the wound or accelerate the regeneration of injured tissues..."...
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What is Crispr Gene Editing? The Complete WIRED Guide

What is Crispr Gene Editing? The Complete WIRED Guide | SynBioFromLeukipposInstitute | Scoop.it
How scientists can repurpose a bacterial immune system to alter DNA, making everything from cheap insulin to extra starchy corn.
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Multiplexed and portable nucleic acid detection platform with Cas13, Cas12a, and Csm6

Rapid detection of nucleic acids is integral for clinical diagnostics and biotechnological applications. We recently developed a platform termed SHERLOCK (specific high-sensitivity enzymatic reporter unlocking) that combines isothermal preamplification with Cas13 to detect single molecules of RNA or DNA. Through characterization of CRISPR enzymology and application development, we report here four advances integrated into SHERLOCK version 2 (SHERLOCKv2) (i) four-channel single-reaction multiplexing with orthogonal CRISPR enzymes; (ii) quantitative measurement of input as low as 2 attomolar; (iii) 3.5-fold increase in signal sensitivity by combining Cas13 with Csm6, an auxiliary CRISPR-associated enzyme; and (iv) lateral-flow readout. SHERLOCKv2 can detect Dengue or Zika virus single-stranded RNA as well as mutations in patient liquid biopsy samples via lateral flow, highlighting its potential as a multiplexable, portable, rapid, and quantitative detection platform of nucleic acids.
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Next-generation diagnostics with CRISPR

Rapid and accurate identification of infectious diseases is essential to optimize clinical care and guide infection control and public health interventions to limit disease spread both in highly specialized medical centers and remote health care settings. The ideal diagnostic test would be inexpensive, accurate, and provide a result rapidly, allowing for point-of-care use on multiple specimen types without need for technical expertise, ancillary equipment, or power. Such a test for highly pathogenic viruses that emerge in remote settings but might spread globally (for example, Ebola virus and Middle East respiratory syndrome coronavirus) would aid in early case detection and isolation, limiting disease spread and facilitating timely care (1). The sentinel discovery that prokaryotes (bacteria and archaea) have heritable adaptive immunity mediated through CRISPR and CRISPR-associated (Cas) proteins has led to transformative advances in molecular biology, most notably in gene editing (2). On pages 436, 444, and 439 of this issue, Chen et al. (3), Myhrvold et al. (4), and Gootenberg et al. (5), respectively, highlight how evolving insights into CRISPR-Cas biology are also revolutionizing the field of molecular diagnostics for infectious diseases, through detection of Zika virus (ZIKV), Dengue virus (DENV), and human papillomavirus (HPV) in human samples, and noninfectious diseases, such as detection of gene mutations in circulating cell-free DNA from lung cancer patients.
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Synthetic biology-based cellular biomedical tattoo for detection of hypercalcemia associated with cancer

Diagnosis marks the beginning of any successful therapy. Because many medical conditions progress asymptomatically over extended periods of time, their timely diagnosis remains difficult, and this adversely affects patient prognosis. Focusing on hypercalcemia associated with cancer, we aimed to develop a synthetic biology-inspired biomedical tattoo using engineered cells that would (i) monitor long-term blood calcium concentration, (ii) detect onset of mild hypercalcemia, and (iii) respond via subcutaneous accumulation of the black pigment melanin to form a visible tattoo. For this purpose, we designed cells containing an ectopically expressed calcium-sensing receptor rewired to a synthetic signaling cascade that activates expression of transgenic tyrosinase, which produces melanin in response to persistently increased blood Ca2+. We confirmed that the melanin-generated color change produced by this biomedical tattoo could be detected with the naked eye and optically quantified. The system was validated in wild-type mice bearing subcutaneously implanted encapsulated engineered cells. All animals inoculated with hypercalcemic breast and colon adenocarcinoma cells developed tattoos, whereas no tattoos were seen in animals inoculated with normocalcemic tumor cells. All tumor-bearing animals remained asymptomatic throughout the 38-day experimental period. Although hypercalcemia is also associated with other pathologies, our findings demonstrate that it is possible to detect hypercalcemia associated with cancer in murine models using this cell-based diagnostic strategy.
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Optogenetics: A Primer for Chemists

The field of optogenetics uses genetically encoded, light-responsive proteins to control physiological processes. This technology has been hailed as the one of the ten big ideas in brain science in the past decade,[1] the breakthrough of the decade,[2] and the method of the year in 2010[3] and again in 2014[4]. The excitement evidenced by these proclamations is confirmed by a couple of impressive numbers. The term "optogenetics" was coined in 2006.[5] As of December 2017, "optogenetics" is found in the title or abstract of almost 1600 currently funded National Institutes of Health grants. In addition, nearly 600 reviews on optogenetics have appeared since 2006, which averages out to approximately one review per week! However, in spite of these impressive numbers, the potential applications and implications of optogenetics are not even close to being fully realized. This is due, in large part, to the challenges associated with the design of optogenetic analogs of endogenous proteins. This review is written from a chemist's perspective, with a focus on the molecular strategies that have been developed for the construction of optogenetic proteins.
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Self-replication of DNA by its encoded proteins in liposome-based synthetic cells

Self-replication of DNA by its encoded proteins in liposome-based synthetic cells | SynBioFromLeukipposInstitute | Scoop.it
Replicating DNA and converting genetic information to protein is a feature of cellular life. Here the authors implement a coupled DNA replication and gene expression system inside vesicles.
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Water-Based Digital Fabrication Platforms : fabrication platform

Water-Based Digital Fabrication Platforms : fabrication platform | SynBioFromLeukipposInstitute | Scoop.it
fabrication platform - In her water-based digital fabrication platform project, architect and professor Neri Oxman embraces water as nature’s architectural tool and...
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neri oxman and MIT develop digitally produced water-based renewable material

neri oxman and MIT develop digitally produced water-based renewable material | SynBioFromLeukipposInstitute | Scoop.it
neri oxman and the MIT mediated matter group have developed a water-based digital fabrication platform using a renewable polymer from the ocean.
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Construction and Integration of a Synthetic MicroRNA Cluster for Multiplex RNA Interference in Mammalian Cells

Methods Mol Biol. 2018;1772:347-359. doi: 10.1007/978-1-4939-7795-6_19.
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At GP-write, scientists take first steps on way to synthetic human genome

At GP-write, scientists take first steps on way to synthetic human genome | SynBioFromLeukipposInstitute | Scoop.it
At the third meeting of GP-write, researchers decide to create virus-resistant human cells...
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Robust Synthetic Circuits for Two-Dimensional Control of Gene Expression in Yeast

Robust Synthetic Circuits for Two-Dimensional Control of Gene Expression in Yeast | SynBioFromLeukipposInstitute | Scoop.it
Robust Synthetic Circuits for Two-Dimensional Control of Gene Expression in Yeast...
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Biology Will Be the Next Great Computing Platform

Biology Will Be the Next Great Computing Platform | SynBioFromLeukipposInstitute | Scoop.it
BIOLOGY WILL BE THE NEXT GREAT COMPUTING PLATFORM
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Amino-acid-encoded biocatalytic self-assembly enables the formation of transient conducting nanostructures

Amino-acid-encoded biocatalytic self-assembly enables the formation of transient conducting nanostructures | SynBioFromLeukipposInstitute | Scoop.it
Aqueous compatible supramolecular materials hold promise for applications in environmental remediation, energy harvesting and biomedicine. One remaining challenge is to actively select a target structure from a multitude of possible options, in response to chemical signals, while maintaining constant, physiological conditions. Here, we demonstrate the use of amino acids to actively decorate a self-assembling core molecule in situ, thereby controlling its amphiphilicity and consequent mode of assembly. The core molecule is the organic semiconductor naphthalene diimide, functionalized with D- and L- tyrosine methyl esters as competing reactive sites. In the presence of α-chymotrypsin and a selected encoding amino acid, kinetic competition between ester hydrolysis and amidation results in covalent or non-covalent amino acid incorporation, and variable supramolecular self-assembly pathways. Taking advantage of the semiconducting nature of the naphthalene diimide core, electronic wires could be formed and subsequently degraded, giving rise to temporally regulated electro-conductivity.
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Field-deployable viral diagnostics using CRISPR-Cas13

Mitigating global infectious disease requires diagnostic tools that are sensitive, specific, and rapidly field deployable. In this study, we demonstrate that the Cas13-based SHERLOCK (specific high-sensitivity enzymatic reporter unlocking) platform can detect Zika virus (ZIKV) and dengue virus (DENV) in patient samples at concentrations as low as 1 copy per microliter. We developed HUDSON (heating unextracted diagnostic samples to obliterate nucleases), a protocol that pairs with SHERLOCK for viral detection directly from bodily fluids, enabling instrument-free DENV detection directly from patient samples in <2 hours. We further demonstrate that SHERLOCK can distinguish the four DENV serotypes, as well as region-specific strains of ZIKV from the 2015–2016 pandemic. Finally, we report the rapid (<1 week) design and testing of instrument-free assays to detect clinically relevant viral single-nucleotide polymorphisms.
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CRISPR-Cas12a target binding unleashes indiscriminate single-stranded DNase activity

Taking CRISPR technology further
CRISPR techniques are allowing the development of technologies for nucleic acid detection (see the Perspective by Chertow). Taking advantages of the distinctive enzymatic properties of CRISPR enzymes, Gootenberg et al. developed an improved nucleic acid detection technology for multiplexed quantitative and highly sensitive detection, combined with lateral flow for visual readout. Myhrvold et al. added a sample preparation protocol to create a field-deployable viral diagnostic platform for rapid detection of specific strains of pathogens in clinical samples. Cas12a (also known as Cpf1), a type V CRISPR protein, cleaves double-stranded DNA and has been adapted for genome editing. Chen et al. discovered that Cas12a also processes single-stranded DNA threading activity. A technology platform based on this activity detected human papillomavirus in patient samples with high sensitivity.
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Computational Re-design of Synthetic Genetic Oscillators for Independent Amplitude and Frequency Modulation

amplitude and period. This need is currently unmet. Here, we demonstrate computationally how two classic genetic oscillators, the dual-feedback oscillator and the repressilator, can be re-designed to provide independent control of amplitude and period and improve tunability-that is, a broad dynamic range of periods and amplitudes accessible through the input "dials." Our approach decouples frequency and amplitude modulation by incorporating an orthogonal "sink module" where the key molecular species are channeled for enzymatic degradation. This sink module maintains fast oscillation cycles while alleviating the translational coupling between the oscillator's transcription factors and output. We characterize the behavior of our re-designed oscillators over a broad range of physiologically reasonable parameters, explain why this facilitates broader function and control, and provide general design principles for building synthetic genetic oscillators that are more precisely controllable.
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Promoters, initiators of transcription and drivers of synthetic biology

Promoters, initiators of transcription and drivers of synthetic biology | SynBioFromLeukipposInstitute | Scoop.it
Promoter characterisation and engineering is a common and arguably useful component of many synthetic biology studies. This post covers some recent articles
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Synthetic biology-based cellular biomedical tattoo for detection of hypercalcemia associated with cancer

Diagnosis marks the beginning of any successful therapy. Because many medical conditions progress asymptomatically over extended periods of time, their timely diagnosis remains difficult, and this adversely affects patient prognosis. Focusing on hypercalcemia associated with cancer, we aimed to develop a synthetic biology-inspired biomedical tattoo using engineered cells that would (i) monitor long-term blood calcium concentration, (ii) detect onset of mild hypercalcemia, and (iii) respond via subcutaneous accumulation of the black pigment melanin to form a visible tattoo. For this purpose, we designed cells containing an ectopically expressed calcium-sensing receptor rewired to a synthetic signaling cascade that activates expression of transgenic tyrosinase, which produces melanin in response to persistently increased blood Ca2+ We confirmed that the melanin-generated color change produced by this biomedical tattoo could be detected with the naked eye and optically quantified. The system was validated in wild-type mice bearing subcutaneously implanted encapsulated engineered cells. All animals inoculated with hypercalcemic breast and colon adenocarcinoma cells developed tattoos, whereas no tattoos were seen in animals inoculated with normocalcemic tumor cells. All tumor-bearing animals remained asymptomatic throughout the 38-day experimental period. Although hypercalcemia is also associated with other pathologies, our findings demonstrate that it is possible to detect hypercalcemia associated with cancer in murine models using this cell-based diagnostic strategy.
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