scholarly journals hei-tag: a highly efficient tag to boost targeted genome editing

2021 ◽  
Author(s):  
Thomas Thumberger ◽  
Tinatini Tavhelidse ◽  
Jose Arturo Gutierrez-Triana ◽  
Rebekka Medert ◽  
Alex Cornean ◽  
...  
Keyword(s):  
Tissue Culture ◽  
Genome Editing ◽  
Nuclear Localization ◽  
High Efficiency ◽  
Basic Research ◽  
Model Systems ◽  
Nuclear Localization Signals ◽  
Highly Efficient ◽  
Cas9 Protein ◽  
Steric Hinderance

Precise, targeted genome editing by CRISPR/Cas9 is key for basic research and translational approaches in model and non-model systems. While active in all species tested so far, editing efficiencies still leave room for improvement. To reach its target, the bacterial Cas9 needs to be efficiently shuttled into the nucleus as attempted by fusion of nuclear localization signals (NLSs) to the Cas9 protein. Additional domains such as FLAG- or myc-tags are added for immediate detection or straight-forward purification. To avoid steric hinderance impacting on activity, amino acid linkers are employed connecting Cas9 and additional domains. We present the 'hei-tag (high efficiency-tag)', boosting the activity of the wide variety of CRISPR/Cas genome editing tools. The addition of the hei-tag to Cas9 or a C-to-T base editor dramatically enhances the respective targeting efficiency in model systems ranging from fish to mammals, including tissue culture applications. This allows to instantly upgrade existing and potentially highly adapted systems as well as establish novel highly efficient tools.

Highly efficient heritable genome editing in wheat using an RNA virus and bypassing tissue culture

2021 ◽  
Author(s):  
Tingdong Li ◽  
Jiacheng Hu ◽  
Yu Sun ◽  
Boshu Li ◽  
Dingliang Zhang ◽  
...  
Keyword(s):  
Tissue Culture ◽  
Genome Editing ◽  
Rna Virus ◽  
Highly Efficient

scholarly journals Highly efficient genome editing of human hematopoietic stem cells via a nano-silicon-blade delivery approach

2017 ◽  
Vol 9 (6) ◽  
pp. 548-554 ◽  
Author(s):  
Yuan Ma ◽  
Xin Han ◽  
Oscar Quintana Bustamante ◽  
Ricardo Bessa de Castro ◽  
Kai Zhang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Genome Editing ◽  
High Efficiency ◽  
Hematopoietic Stem ◽  
Highly Efficient ◽  
Nano Silicon ◽  
Delivery Approach

We provided a nano-blade chip for HSCs specific delivery with the properties of rapid, high efficiency and harmless.

scholarly journals Various Aspects of a Gene Editing System—CRISPR–Cas9

2020 ◽  
Vol 21 (24) ◽  
pp. 9604
Author(s):  
Edyta Janik ◽  
Marcin Niemcewicz ◽  
Michal Ceremuga ◽  
Lukasz Krzowski ◽  
Joanna Saluk-Bijak ◽  
...  
Keyword(s):  
Genome Editing ◽  
Gene Editing ◽  
Genome Engineering ◽  
High Efficiency ◽  
Adaptive Immune System ◽  
Zinc Finger Nucleases ◽  
Transcription Activator ◽  
Guide Rna ◽  
Adaptive Immune ◽  
Cas9 Protein

The discovery of clustered, regularly interspaced short palindromic repeats (CRISPR) and their cooperation with CRISPR-associated (Cas) genes is one of the greatest advances of the century and has marked their application as a powerful genome engineering tool. The CRISPR–Cas system was discovered as a part of the adaptive immune system in bacteria and archaea to defend from plasmids and phages. CRISPR has been found to be an advanced alternative to zinc-finger nucleases (ZFN) and transcription activator-like effector nucleases (TALEN) for gene editing and regulation, as the CRISPR–Cas9 protein remains the same for various gene targets and just a short guide RNA sequence needs to be altered to redirect the site-specific cleavage. Due to its high efficiency and precision, the Cas9 protein derived from the type II CRISPR system has been found to have applications in many fields of science. Although CRISPR–Cas9 allows easy genome editing and has a number of benefits, we should not ignore the important ethical and biosafety issues. Moreover, any tool that has great potential and offers significant capabilities carries a level of risk of being used for non-legal purposes. In this review, we present a brief history and mechanism of the CRISPR–Cas9 system. We also describe on the applications of this technology in gene regulation and genome editing; the treatment of cancer and other diseases; and limitations and concerns of the use of CRISPR–Cas9.

scholarly journals NT-CRISPR: Combining natural transformation and CRISPR/Cas9 counterselection for markerless and scarless genome editing in Vibrio natriegens

2021 ◽  
Author(s):  
Daniel Stukenberg ◽  
Josef Hoff ◽  
Anna Faber ◽  
Anke Becker
Keyword(s):  
Genome Editing ◽  
High Efficiency ◽  
Natural Transformation ◽  
Single Base ◽  
Highly Efficient ◽  
Target Spectrum ◽  
A Genome ◽  
Wide Range ◽  
Resistance Markers ◽  
Type Sequence

The fast-growing bacterium Vibrio natriegens has recently gained increasing attention as a novel chassis organism for a wide range of projects. To fully harness the potential of this fascinating bacterium, convenient and highly efficient genome editing methods are indispensable to create novel strains, tailored for specific applications. V. natriegens is able to take up free DNA and incorporate it into its genome by homologous recombination. This process, called natural transformation, was tamed for genome editing. It displays a high efficiency and is able to mediate uptake of multiple DNA fragments, thereby allowing multiple simultaneous edits. Here, we describe NT-CRISPR, a combination of natural transformation with CRISPR/Cas9 counterselection. In two temporally distinct steps, we first performed a genome edit by natural transformation and second, induced CRISPR/Cas9, targeting the wild type sequence, leading to death of non-edited cells. Through highly efficient cell killing with efficiencies of up to 99.999 %, integration of antibiotic resistance markers became dispensable and thus enabled scarless and markerless edits with single-base precision. We used NT-CRISPR for deletions, integrations and single-base modifications with editing efficiencies of up to 100 % and further demonstrated its applicability for the simultaneous deletion of multiple chromosomal regions. Lastly, we demonstrated that the near PAM-less Cas9 variant SpG Cas9 is compatible with NT-CRISPR and thereby massively broadens the target spectrum.

scholarly journals Rapid target validation in a Cas9-inducible hiPSC derived kidney model

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yasaman Shamshirgaran ◽  
Anna Jonebring ◽  
Anna Svensson ◽  
Isabelle Leefa ◽  
Mohammad Bohlooly-Y ◽  
...  
Keyword(s):  
High Efficiency ◽  
Disease Modeling ◽  
Genetic Manipulation ◽  
Disease Model ◽  
Genetically Engineered ◽  
Model Systems ◽  
Target Validation ◽  
Direct Differentiation ◽  
Kidney Organoids

AbstractRecent advances in induced pluripotent stem cells (iPSCs), genome editing technologies and 3D organoid model systems highlight opportunities to develop new in vitro human disease models to serve drug discovery programs. An ideal disease model would accurately recapitulate the relevant disease phenotype and provide a scalable platform for drug and genetic screening studies. Kidney organoids offer a high cellular complexity that may provide greater insights than conventional single-cell type cell culture models. However, genetic manipulation of the kidney organoids requires prior generation of genetically modified clonal lines, which is a time and labor consuming procedure. Here, we present a methodology for direct differentiation of the CRISPR-targeted cell pools, using a doxycycline-inducible Cas9 expressing hiPSC line for high efficiency editing to eliminate the laborious clonal line generation steps. We demonstrate the versatile use of genetically engineered kidney organoids by targeting the autosomal dominant polycystic kidney disease (ADPKD) genes: PKD1 and PKD2. Direct differentiation of the respective knockout pool populations into kidney organoids resulted in the formation of cyst-like structures in the tubular compartment. Our findings demonstrated that we can achieve > 80% editing efficiency in the iPSC pool population which resulted in a reliable 3D organoid model of ADPKD. The described methodology may provide a platform for rapid target validation in the context of disease modeling.

scholarly journals A self-boosting microwave plasma strategy tuned by air pressure for the highly efficient and controllable surface modification of carbon

RSC Advances ◽  
2021 ◽  
Vol 11 (17) ◽  
pp. 9955-9963
Author(s):  
Yanjing Liu ◽  
Jiawei He ◽  
Bing Zhang ◽  
Huacheng Zhu ◽  
Yang Yang ◽  
...  
Keyword(s):  
Surface Modification ◽  
Carbon Fiber ◽  
High Efficiency ◽  
Microwave Plasma ◽  
Air Pressure ◽  
Air Plasma ◽  
Highly Efficient ◽  
Carbon Fiber Cloth

Microwave enabled air plasma was boosted by a carbon fiber cloth (CFC) and used for the high-efficiency surface modification of the CFC, yielding CFCs with tunable contents of oxygen and each O-containing group.

Unprecedentedly high efficiency for photocatalytic conversion of methane to methanol over Au–Pd/TiO2 – what is the role of each component in the system?

2021 ◽  
Author(s):  
Xiaojiao Cai ◽  
Siyuan Fang ◽  
Yun Hang Hu
Keyword(s):  
Methane Conversion ◽  
High Efficiency ◽  
Mild Conditions ◽  
Highly Efficient ◽  
System P ◽  
Conversion Of Methane

Direct and highly efficient methane conversion to methanol under mild conditions is achieved via photocatalysis over Au–Pd/TiO2.

Maize tissue culture, transformation, and genome editing

2021 ◽  
Author(s):  
Albert P. Kausch ◽  
Kimberly Nelson-Vasilchik ◽  
Michael Tilelli ◽  
Joel P. Hague
Keyword(s):  
Tissue Culture ◽  
Genome Editing ◽  
Culture Transformation ◽  
Maize Tissue Culture

scholarly journals A general approach to high-efficiency perovskite solar cells by any antisolvent

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Alexander D. Taylor ◽  
Qing Sun ◽  
Katelyn P. Goetz ◽  
Qingzhi An ◽  
Tim Schramm ◽  
...  
Keyword(s):  
Solar Cells ◽  
High Efficiency ◽  
Perovskite Solar Cells ◽  
Precursor Solution ◽  
Microstructural Characterization ◽  
Application Rate ◽  
Perovskite Layer ◽  
Highly Efficient ◽  
Application Procedure ◽  
Wide Range

AbstractDeposition of perovskite films by antisolvent engineering is a highly common method employed in perovskite photovoltaics research. Herein, we report on a general method that allows for the fabrication of highly efficient perovskite solar cells by any antisolvent via manipulation of the antisolvent application rate. Through detailed structural, compositional, and microstructural characterization of perovskite layers fabricated by 14 different antisolvents, we identify two key factors that influence the quality of the perovskite layer: the solubility of the organic precursors in the antisolvent and its miscibility with the host solvent(s) of the perovskite precursor solution, which combine to produce rate-dependent behavior during the antisolvent application step. Leveraging this, we produce devices with power conversion efficiencies (PCEs) that exceed 21% using a wide range of antisolvents. Moreover, we demonstrate that employing the optimal antisolvent application procedure allows for highly efficient solar cells to be fabricated from a broad range of precursor stoichiometries.

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