Tag: crRNA

Biochemical characterization of the two novel mgCas12a proteins from the human gut metagenome

Discovery of 11 new Cas12a candidates from a human gut microbiome database To discover new members of the Cas12a protein family, we built a pipeline that searches and sorts out new candidates from the National Center for Biotechnological Information (NCBI) human metagenome database. Using this search pipeline (Fig. 1a), we initially…

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Extra-hematopoietic immunomodulatory role of the guanine-exchange factor DOCK2

Cell isolation, reprogramming and culture Approval was obtained for human cell and tissue sample collection and genetic reprogramming from the Institutional Review Board (protocols 19–252, 18–243, 21–060, 19–284 and 415-E/1776/4-2014, Ethics Committee of the province of Salzburg). Adult samples were collected in accordance with the Declaration of Helsinki after written…

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CVC Files Appeal Brief in Interference No. 106,115 | McDonnell Boehnen Hulbert & Berghoff LLP

The Patent Trial and Appeal Board (like its predecessor, the Board of Patent Appeals and Interferences or BPAI) occasionally renders an opinion having the tendency to raise an eyebrow or two, which on occasion has led the Federal Circuit, like its predecessor the Court of Claims and Patent Appeals, to…

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Multiplexed base editing through Cas12a variant-mediated cytosine and adenine base editors

Robust C-to-T conversion by Cas12a variant-mediated CBEs for non-canonical PAMs We initially attempted to screen for optimal cytosine deaminases that confer powerful C-to-T editing for Cas12a-mediated CBEs. Seven dCas12a-CBEs fused with individual cytosine deaminases1,2,16,20,21,22,24,25,26 were compared in HEK293T cells and porcine embryos at sites with TTTV PAM sequences. The dCas12a-A3A…

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731 Type VI A Cas13a In Cas13a direct repeat region of crRNA is composed of 5 6

7.3.1Type VI-A (Cas13a)In Cas13a, direct repeat region of crRNA is composed of 5–6 bp stem using Watsonand Crick base pairing with an unpaired 2 nt bulge (AA or AC) present at 30end ofstem, while loop region of stem contains 7–9 nt depending upon homologs ofCas13a. In addition, an 8–12 nt…

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How is RNA used in CRISPR and mRNA vaccines?

Why is RNA important in immunity? All organisms, from single-celled bacteria to blue whales, have a genome that’s made of deoxyribonucleic acid, or DNA. When it comes to viruses, many use single-stranded ribonucleic acid (RNA). However, certain species (such as rotavirus, which causes gastroenteritis) have a double-stranded RNA genome. The…

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Inferring and perturbing cell fate regulomes in human brain organoids

Experimental methods Stem cell and organoid culture We used six human iPS cell lines (Hoik1, Wibj2, Kucg2 from the HipSci resource47; 409B2 from the RIKEN BRC cell bank; 01F49i-N-B7 (B7) from Institute of Molecular and Clinical Ophthalmology Basel; and WTC from the Allen Institute) and three human ES cell lines (H1-PAX6YFP…

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Dynamic mechanisms of CRISPR interference by Escherichia coli CRISPR-Cas3

In vitro reconstitution of Escherichia coli CRISPR-Cas3 interference E. coli CRISPR-Cas3 is generally well-characterized type I CRISPR complexes in vitro and in vivo32,33,37,38. However, recombinant EcoCas3 protein is difficult to purify because of poor solubility and propensity to aggregate at 37 °C25,26,30,39. Co-expression of HtpG chaperon40 and/or low temperature growth at…

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Multidrug resistance microbial therapy | VMRR

Introduction Antimicrobials are the most important and useful therapeutic discovery in the history of medicine. It allows living beings to survive the microbial disease, enhances invasive surgical procedures, ensures animal health, and protects the food chain since 1928 after the discovery of penicillin. However, the indiscriminate use of antibiotics and…

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New dual-plasmid editing system for DNA-based information rewriting in vivo

DNA-based information is a new interdisciplinary field linking information technology and biotechnology. The field hopes to meet the enormous need for long-term data storage by using DNA as an information storage medium. Despite DNA’s promise of strong stability, high storage density and low maintenance cost, however, researchers face problems accurately…

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Amount of Cas9 protein introduced into mouse embryos via electroporation affects the genome-editing rate

doi: 10.1262/jrd.2022-067. Online ahead of print. Affiliations Expand Affiliations 1 Division of Science and Engineering, Graduate School of Arts and Science, Iwate University, Iwate 020-8551, Japan. 2 Integrated DNA Technologies, Inc., Coralville, IA 52241, U.S.A. 3 Department of Chemistry and Biological Sciences, Faculty of Science and Engineering, Iwate University, Iwate 020-8551,…

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CRISPR Gene Detection and Diagnostic Industry 2022 Market Size, Share and Growth, Global Segments Analysis and Dynamic Research Report 2029

The Wide Ranging CRISPR Gene Detection and Diagnostic Business Report encompasses a thorough study of current situation of the global market along with several market dynamics. The market research report is a demonstrated source of information which offers a telescopic view of the current market trends, size, share, growth, demand,…

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A closed Candidatus Odinarchaeum chromosome exposes Asgard archaeal viruses

Zaremba-Niedzwiedzka, K. et al. Asgard archaea illuminate the origin of eukaryotic cellular complexity. Nature 541, 353–358 (2017). CAS  PubMed  Article  Google Scholar  Williams, T. A., Cox, C. J., Foster, P. G., Szöllősi, G. J. & Embley, T. M. Phylogenomics provides robust support for a two-domains tree of life. Nat. Ecol….

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A highly sensitive fluorescent aptasensor for detection of prostate specific antigen based on the integration of a DNA structure and CRISPR-Cas12a

. 2022 Aug 1;1219:340031. doi: 10.1016/j.aca.2022.340031. Epub 2022 Jun 3. Affiliations Expand Affiliations 1 Targeted Drug Delivery Research Center, Pharmaceutical Technology Institute, Mashhad University of Medical Sciences, Mashhad, Iran; Department of Pharmaceutical Biotechnology, School of Pharmacy, Mashhad University of Medical Sciences, Mashhad, Iran. 2 Pharmaceutical Research Center, Pharmaceutical Technology Institute,…

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Bio-informatic analysis of CRISPR protospacer adjacent motifs (PAMs) in T4 genome | BMC Genomic Data

The clustered regularly interspaced short palindromic repeats-Cas (CRISPR-Cas) defensive system is an acquired mechanism in prokaryotes which is analogous to RNAi in eukaryotes [1]. The CRISPR machinery is a complex immune strategy that is continually evolving in the bacterial genome to accommodate the rapid changes in the nucleic acids of…

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Frontiers | A Mutated Nme1Cas9 Is a Functional Alternative RNase to Both LwaCas13a and RfxCas13d in the Yeast S. cerevisiae

Introduction The clustered regularly interspaced short palindromic repeats (CRISPR)–CRISPR-associated (Cas) protein systems naturally exist in prokaryotes. They are RNA-mediated defense mechanisms to protect bacteria and archaea from invading nucleic acids (Barrangou et al., 2007). CRISPR–Cas systems are classified into two classes (1 and 2). Class 2 CRISPR–Cas require a single…

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Specific High-Sensitivity Enzymatic Molecular Detection System Termed RPA-Based CRISPR-Cas13a for Duck Tembusu Virus Diagnostics

journal contribution posted on 19.05.2022, 07:04 by Dalin He, Gang Liu, Jing Yang, Xiaoning Jiang, Hongzhi Wang, Yaru Fan, Shangyu Gong, Feng Wei, Youxiang Diao, Yi Tang In China, drastic losses in the economy have been caused by the Tembusu virus (TMUV), the causative agent of the egg-drop syndrome, to…

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A Highly Sensitive and Specific Detection Method for Mycobacterium tuberculosis Fluoroquinolone Resistance Mutations Utilizing the CRISPR-Cas13a System

Objectives CRISPR-Cas13a system-based nucleic acid detection methods are reported to have rapid and sensitive DNA detection. However, the screening strategy for crRNAs that enables CRISPR-Cas13a single-base resolution DNA detection of human pathogens remains unclear. Methods A combined rational design and target mutation-anchoring CRISPR RNA (crRNA) screening strategy was proposed. Results…

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WGS Facilitates Gene Editing System Upgrade

Researchers at the Korean Institute of Life Sciences and Technology engineered an efficient, miniaturized CRISPR-Cas gene-editing system that may be more easily packed into vectors for clinical applications. Their system employs the Cas variant Cas12f1 with a guide RNA (gRNA) remodeled to mitigate off-target effects, a design that could potentially…

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Multiplexed genome regulation in vivo with hyper-efficient Cas12a

. 2022 Apr;24(4):590-600. doi: 10.1038/s41556-022-00870-7. Epub 2022 Apr 12. Lucie Y Guo #  1   2 , Jing Bian #  3 , Alexander E Davis  4 , Pingting Liu  4 , Hannah R Kempton  3 , Xiaowei Zhang  3 , Augustine Chemparathy  3 , Baokun Gu  3 , Xueqiu Lin  3 , Draven A Rane  3 , Xiaoshu Xu  3 , Ryan M…

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RCSB PDB – 7V94: Cryo-EM structure of the Cas12c2-sgRNA-target DNA ternary complex

RNA-guided CRISPR-Cas nucleases are widely used as versatile genome-engineering tools. Recent studies identified functionally divergent type V Cas12 family enzymes. Among them, Cas12c2 binds a CRISPR RNA (crRNA) and a trans-activating crRNA (tracrRNA) and recognizes double-stranded DNA targets with a short TN PAM … RNA-guided CRISPR-Cas nucleases are widely used…

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Multiplexed genome regulation in vivo with hyper-efficient Cas12a,Nature Cell Biology

Multiplexed modulation of endogenous genes is crucial for sophisticated gene therapy and cell engineering. CRISPR–Cas12a systems enable versatile multiple-genomic-loci targeting by processing numerous CRISPR RNAs (crRNAs) from a single transcript; however, their low efficiency has hindered in vivo applications. Through structure-guided protein engineering, we developed a hyper-efficient Lachnospiraceae bacterium Cas12a…

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Sigma-Aldrich Files Opposition to CVC Substantive Preliminary Motion No. 1 to be Accorded Priority Benefit | McDonnell Boehnen Hulbert & Berghoff LLP

On February 18th, Sigma-Aldrich filed its Opposition to Junior Party’s (the University of California, Berkeley; the University of Vienna; and Emmanuelle Charpentier; collectively, “CVC”) Substantive Preliminary Motion No. 1 in Interference No. 106,132, asking the Patent Trial and Appeal Board for benefit of priority to U.S. Provisional Application No. 61/652,086,…

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Cho Cell Line Development & Engineering: Impact On Biopharmaceutical Production: Essay Example, 2571 words

1. Introduction The approval of Chinese hamster ovary (CHO) cell cultures for the manufacturing of protein therapeutic products modernized conventional medicine. CHO cells remained the workhorse for the production of monoclonal antibodies (mAbs) for the last at least 20 years for different reasons. First, CHO cells are easily adapting and…

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Development of Cas12a-Based Cell-Free Small-Molecule Biosensors via Allosteric Regulation of CRISPR Array Expression

In nature, microbes have evolved different systems to sense external stimuli. Synthetic biology approaches (1) repurpose these systems as biosensors to specifically and sensitively detect various targets of interest. Although various highly sensitive and specific laboratory-based analytical methods (including high-performance liquid chromatography and mass spectrometry) can detect small-molecule targets, they…

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CRISPR/Cas12a-based technology: A powerful tool for biosensing in food safety

Background: In the context of the current pandemic caused by the novel coronavirus, molecular detection is not limited to the clinical laboratory, but also faces the challenge of the complex and variable real-time detection fields. A series of novel coronavirus events were detected in the process of food cold chain…

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A combinatorial CRISPR-Cas12a attack on HIV DNA: Molecular Therapy

CRISPR-Cas12a is an alternative class 2 gene-editing tool that may cause less off-target effects than the original Cas9 system. We have previously demonstrated that Cas12a attack with a single CRISPR RNA (crRNA) can neutralize all infectious HIV in an infected T cell line in cell culture. However, we demonstrated that…

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LAMP-CRISPR-Cas12a-lateral flow immunochromatographic strip | IDR

Introduction About 700,000 people die from “superbugs” globally every year. Antibiotic abuse is the culprit of extensive spread of superbugs. Currently, carbapenemase-producing organisms are among the most important pathogens of hospital infections. Especially the plasmid-mediated highly transmissible carbapenem-resistant Enterobacterales (CRE) has become an important public health issue of global concern….

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Highly Efficient Genome Editing in Plant Protoplasts by Ribonucleoprotein Delivery of CRISPR-Cas12a Nucleases

Review doi: 10.3389/fgeed.2022.780238. eCollection 2022. Affiliations Expand Affiliations 1 Department of Plant Science and Landscape Architecture, University of Maryland, College Park, College Park, MD, United States. 2 College of Agriculture, Nanjing Agricultural University, Nanjing, China. 3 Integrated DNA Technologies, Coralville, IA, United States. 4 U.S. Horticultural Research Laboratory, USDA-Agricultural Research…

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The comparisonof the function of crRNA (cr-ribonucleic acid) and tracrRNA (trans-activating crRNA) in the CRISPR-Cas(clustered regularly interspaced short palindromic repeats-CRISPR associated protein 9)system. Introduction: The CRISPR-Cas system is a crucial defense system against foreign invaders, like bacteriophage, transposons, and plasmids in many prokaryotes. There are five genes present on it: tracr, Cas1, Crispr, Cas2, Cas9 on the CRISPR. The tracrRNA and the crRNA have a guiding function in the expression phase of the defense mechanism.

Chapter 17, Problem 11CONQ To review: The comparisonof the function of crRNA (cr-ribonucleic acid) and tracrRNA (trans-activating crRNA) in the CRISPR-Cas(clustered regularly interspaced short palindromic repeats-CRISPR associated protein 9)system. Introduction: The CRISPR-Cas system is a crucial defense system against foreign invaders, like bacteriophage, transposons, and plasmids in many prokaryotes….

Continue Reading The comparisonof the function of crRNA (cr-ribonucleic acid) and tracrRNA (trans-activating crRNA) in the CRISPR-Cas(clustered regularly interspaced short palindromic repeats-CRISPR associated protein 9)system. Introduction: The CRISPR-Cas system is a crucial defense system against foreign invaders, like bacteriophage, transposons, and plasmids in many prokaryotes. There are five genes present on it: tracr, Cas1, Crispr, Cas2, Cas9 on the CRISPR. The tracrRNA and the crRNA have a guiding function in the expression phase of the defense mechanism.

A Simple and Highly Sensitive Naked-Eye Analysis of EGFR 19del via CRISPR/Cas12a Triggered No-Nonspecific Nucleic Acid Amplification

The mutation status of epidermal growth factor receptor (EGFR) exon 19 is of great importance for predicting sensitivity to tyrosine kinase inhibitors (TKIs) in the treatment of non-small-cell lung cancer (NSCLC). However, the development of simple, sensitive, and no-nonspecific amplification platforms for EGFR 19del detection in NSCLC remains a challenge….

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Combining functional hairpin probes with disordered cleavage of CRISPR/Cas12a protease to screen for B lymphocytic leukemia

Available online 4 January 2022, 113941 doi.org/10.1016/j.bios.2021.113941Get rights and content Highlights • The sensor based on the label-free functional hairpin probe and the disordered cleavage of CRISPR/Cas12a was successfully constructed, and it can also accurately distinguish target and non-target genes successfully used for the detection of the Pax-5a gene. •…

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PTAB Sets Motions and Times in Broad vs. Sigma Interference No. 106,133 | McDonnell Boehnen Hulbert & Berghoff LLP

Following a telephone conference held on August 16th (a transcript of which can be found here) between the Board and representatives of Junior Party the Broad Institute, Harvard University, and MIT (collectively, Broad) and Senior Party Sigma-Aldrich, the Board issued its Order on September 20th authorizing motions and setting times…

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Efficient target cleavage by Type V Cas12a effectors programmed with split CRISPR RNA

doi: 10.1093/nar/gkab1227. Online ahead of print. Regina Shebanova  1 , Natalia Nikitchina  1   2 , Nikita Shebanov  1 , Vladimir Mekler  3 , Konstantin Kuznedelov  3 , Egor Ulashchik  4 , Ruslan Vasilev  5   6 , Olga Sharko  4 , Vadim Shmanai  4 , Ivan Tarassov  2 , Konstantin Severinov  1   3   7 , Nina Entelis  2…

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CRISPR/Cas12a Powered DNA Framework-Supported Electrochemical Biosensing Platform for Ultrasensitive Nucleic Acid Analysis

Nucleic acid analysis using ultrasensitive and simple methods is critically important for the early-stage diagnosis and treatment of diseases. The CRISPR/Cas proteins, guided by a single-stranded RNA have shown incredible capability for sequence-specific targeting and detection. Herein, in order to improve and expand the application of CRISPR/Cas technology to the…

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Sigma-Aldrich and Broad Propose Preliminary Motions in Recent CRISPR Interference No. 106,133 | McDonnell Boehnen Hulbert & Berghoff LLP

The parties in Interference No. 106,133, namely Senior Party Sigma-Aldrich and Junior Party the Broad Institute, Harvard University, and MIT (collectively, “Broad”), filed their respective lists of proposed preliminary motions four days prior to their August 3rd teleconference with the Board to present their arguments for the Board to grant…

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Sigma-Aldrich Joins the CRISPR Interference Fray | McDonnell Boehnen Hulbert & Berghoff LLP

On June 21st,* the Patent Trial and Appeal Board declared two new interferences involving CRISPR technology.  The first, Interference No. 106,132, named Sigma-Aldrich as Senior Party and the University of California/Berkeley, the University of Vienna, and Emmanuelle Charpentier (collectively, “CVC”) as Junior Party, while the second, Interference No. 106,133 named…

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Recombinase Polymerase Amplification/Cas12a-Based Identification of Xanthomonas arboricola pv. pruni on Peach

doi: 10.3389/fpls.2021.740177. eCollection 2021. Affiliations Expand Affiliations 1 Key Laboratory of Horticultural Plant Biology, Ministry of Education, Huazhong Agricultural University, Wuhan, China. 2 Hubei Key Laboratory of Plant Pathology, Huazhong Agricultural University, Wuhan, China. Free PMC article Item in Clipboard Mei Luo et al. Front Plant Sci. 2021. Free PMC article…

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CVC Files Opposition to ToolGen Substantive Motion No. 1 | McDonnell Boehnen Hulbert & Berghoff LLP

On July 15th, Junior Party the University of California/Berkeley, the University of Vienna, and Emmanuelle Charpentier (collectively, “CVC”) filed its Opposition to Senior Party ToolGen’s Substantive Motion No. 1 for benefit of priority to U.S. Provisional Application No. 61/837,481, filed June 20, 2013 (“P3” or “ToolGen P3”), or alternatively, International…

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New Techniques and Complex Models

CRISPR systems that rely on inactivated Cas enzymes—that is, dead Cas (dCas) enzymes—never looked more alive. They harness the targeting power associated with CRISPR—but not the double-strand cuts. As such, they give researchers new ways to interrogate and manipulate gene function. Possibilities include CRISPR interference (CRISPRi) and CRISPR activation (CRISPRa)…

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ToolGen Files Opposition to CVC Substantive Preliminary Motion No. 3 to Add Claims in ToolGen Patent | McDonnell Boehnen Hulbert & Berghoff LLP

On May 20th, Junior Party the University of California, Berkeley; the University of Vienna; and Emmanuelle Charpentier (collectively, “CVC”) filed its Substantive Preliminary Motion No. 3 in Interference No. 106,127 (which names ToolGen as Senior Party), asking the Patent Trial and Appeal Board to add claims in ToolGen’s U.S. Patent…

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CRISPR Applications in Medicine Depend on Minimizing Off-Target Editing

By Rolf Turk, PhD Genome editing technologies, such as CRISPR, are being applied to better understand basic biological systems as well as to research new kinds of gene and cell therapies. The CRISPR-Cas9 system comprises a Cas9 endonuclease protein that forms a complex with a guide RNA (gRNA) molecule, which…

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Controlling CRISPR | Optics & Photonics News

Light-based approaches for steering CRISPR–Cas9, the powerful Nobel Prize–winning genome-editing toolkit, promise to boost the technique’s precision and specificity—and might help bring it into clinical use. [Getty Images] In early 2011, two scientists walked the cobblestone streets of Old San Juan, the historic district of Puerto Rico’s capital. Recently introduced…

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CRISPR: Guide to gRNA design

Introduction to CRISPR in SnapGene Genome editing technology has been evolving for many years. The Holy Grail of genome engineering has always been to introduce a specific genetic change that affects only the genomic target and leaves no undesired changes in the DNA. The discovery and application of the bacterial…

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