Correct Copy Research Articles

CRISPR/Cas9-Based Disease Modeling and Functional Correction of Interleukin 7 Receptor Alpha Severe Combined Immunodeficiency in T-Lymphocytes and Hematopoietic Stem Cells.

Interleukin 7 Receptor alpha Severe Combined Immunodeficiency (IL7R-SCID) is a life-threatening disorder caused by homozygous mutations in the IL7RA gene. Defective IL7R expression in humans hampers T cell precursors' proliferation and differentiation during lymphopoiesis resulting in the absence of T cells in newborns, who succumb to severe infections and death early after birth. Previous attempts to tackle IL7R-SCID by viral gene therapy have shown that unregulated IL7R expression predisposes to leukemia, suggesting the application of targeted gene editing to insert a correct copy of the IL7RA gene in its genomic locus and mediate its physiological expression as a more feasible therapeutic approach. To this aim, we have first developed a CRISPR/Cas9-based IL7R-SCID disease modeling system that recapitulates the disease phenotype in primary human T cells and hematopoietic stem and progenitor cells (HSPCs). Then, we have designed a knockin strategy that targets IL7RA exon 1 and introduces through homology-directed repair a corrective, promoterless IL7RA cDNA followed by a reporter cassette through AAV6 transduction. Targeted integration of the corrective cassette in primary T cells restored IL7R expression and rescued functional downstream IL7R signaling. When applied to HSPCs further induced to differentiate into T cells in an Artificial Thymic Organoid system, our gene editing strategy overcame the T cell developmental block observed in IL7R-SCID patients, while promoting full maturation of T cells with physiological and developmentally regulated IL7R expression. Finally, genotoxicity assessment of the CRISPR/Cas9 platform in HSPCs using biased and unbiased technologies confirmed the safety of the strategy, paving the way for a new, efficient, and safe therapeutic option for IL7R-SCID patients.

Using copies can improve precision in continuous-time quantum computing

In the quantum optimisation setting, we build on a scheme introduced by Young et al (2013 Phys. Rev. A 88 062314), where physical qubits in multiple copies of a problem encoded into an Ising spin Hamiltonian are linked together to increase the logical system’s robustness to error. We introduce several innovations that improve the error suppression of this scheme under a special model of control noise, designed to understand how limited precision could be overcome. First, we note that only one copy needs to be correct by the end of the computation, since solution quality can be checked efficiently. Second, we find that ferromagnetic links do not generally help in this ‘one correct copy’ setting, but anti-ferromagnetic links do help on average, by suppressing the chance of the same error being present on all of the copies. Third, we find that minimum-strength anti-ferromagnetic links perform best, by counteracting the spin-flips induced by the errors. We have numerically tested our innovations on small instances of spin glasses from Callison et al (2019 New J. Phys. 21 123022), and we find improved error tolerance for three or more copies in configurations that include frustration. Interpreted as an effective precision increase, we obtain several extra bits of precision on average for three copies connected in a triangle. This provides proof-of-concept of a method for scaling quantum annealing beyond the precision limits of hardware, a step towards fault tolerance in this setting.

The steep uphill path leading to ex vivo gene therapy for genodermatoses.

Cell therapy, gene therapy, and tissue engineering have the potential to revolutionize the field of regenerative medicine. In particular, gene therapy is understood as the therapeutical correction of mutated genes by addition of a correct copy of the gene or site-specific gene modifications. Gene correction of somatic stem cells sustaining renewing tissues is critical to ensure long-term clinical success of ex vivo gene therapy. To date, remarkable clinical outcomes arose from combined ex vivo cell and gene therapy of different genetic diseases, such as immunodeficiencies and genodermatoses. Despite the efforts of researchers around the world, only a few of these advanced approaches have yet made it to routine therapy. In fact, gene therapy poses one of the greatest technical challenges in modern medicine, spanning safety and efficacy issues, regulatory constraints, registration and market access, all of which need to be addressed to make the therapy available to patients with rare disease. In this review, we survey some of the main challenges in the development of combined cell and gene therapy of genetic skin diseases.

Lectin Staining for the Detection of In Vitro Sialic Acid Alterations

GNE Myopathy (GNEM) is a rare disease caused by a mutation in the UDP‐N‐acetylglucosamine 2‐epimerase/N‐acetylmannosamine kinase (GNE) gene, which catalyzes sialic acid synthesis. GNEM results in decreased sialic acid and loss of strength in the muscles due to an unclear mechanism. Currently, there is no available treatment for GNEM. However, gene therapy approaches involving delivering a corrected copy of the GNE gene into GNEM patient muscle are currently being developed. A robust biomarker is needed to assess the efficacy of such a therapy. Here, we assess the ability of a panel of lectins, which are glycan‐binding proteins, to reflect changes in sialic acid levels in vitro that occur when sialic acid is enzymatically removed via sialidase or when its production is impaired due to mutations in the GNEgene. This work probes the ability of these lectins to reflect changes in sialic acid levels, which can inform biomarker development for GNE myopathy therapies.

Corporate Governance And Its Effect On Professional Performance In Palestinian Private Universities, In Light Of Quality, Accreditation, And Classification Requirements

Corporate governance has become a subject of heightened importance and attention in government policy circles, academia, and the popular press throughout all over the world. The purpose of the study is to examine the actual status of corporate governance principles in Palestinian private universities at Gaza Strip Governorates, in order to propose a framework of governance for these universities, in light of quality, accreditation and classification requirements for Palestinian universities issued by Palestinian Ministry of Higher Education. In Gaza Strip Governorates there are four private universities, the study covers all these four universities. To achieve the study objectives, the descriptive analytical approach was implemented. The study population includes all the 458, stockholders, academic and administrative fixed contract employees at the four private universities in Gaza Strip. the sample was a random sample of 200 employees (44% of the population). A special questionnaire was prepared and distributed for the sample, 182 completed and correct questionnaire copies were collected (91% of total sample). Data was collected using Statistical Package for the Social Sciences SPSS program. The main finding of the study demonstrates that there is a significant relationship between private universities performance and the application of corporate governance principles, quality and accreditation requirements and classification requirements of Ministry of Higher Education (MOHE), and applied corporate governance framework, and there is a middle level of adequacy of corporate governance principles adequacy at private universities in Gaza Governorates. These findings reflect the actual situation of all Palestinian private universities. Based on the study results, the researchers recommend to build national standards for university governance, aiming to accelerate the issuance of a guide to the governance of private universities. Also, promoting cultural and knowledge awareness regarding the governance of private universities. These recommendations will help Palestinian private universities to achieve higher performance quality in both academic and managerial aspects, this will improve the outcomes of these universities and will be reflected in the capabilities of their graduate students.

Selection Combining-based Packet Combining Schemes for Multi-path Diversity System

In multi-path diversity system, the receiver receives several signals bearing the same information, through distinct and independent fading channels. The packets traversing through independent and distinct paths would undergo different degree of fading. Thus, even if the copy of the packet is distorted badly in one path, the copy received from other paths might be lesser distorted; so by combining the copies of the packet, the correct copy of the packet could be retrieved. In this paper, selection combining-based packet combining scheme for multi-path diversity system have been proposed. In the proposed scheme, a packet is transmitted to the receiver via $$m$$ different paths, where $$m \ge 2$$ . At the receiver, erroneously received copies of the packet are combined to recover the correct copy of the packet. The proposed selection combining-based packet combining scheme is analyzed and studied using simulation on MatLab. It was observed experimentally that proposed scheme has superior performance than that of conventional packet combining schemes. The superior performance of the proposed scheme with selection combining scheme can be attributed to diversity achieved using multi-path transmission.

Constitutive Model of Erythrocyte Membranes with Distributions of Spectrin Orientations and Lengths

We present an analytical hyperelastic constitutive model of the red blood cell (erythrocyte) membrane based on recently improved characterizations of density and microscopic structure of its spectrin network from proteomics and cryo-electron tomography. The model includes distributions of both orientations and natural lengths of spectrin and updated copy numbers of proteins. By applying finite deformation to the spectrin network, we obtain the total free energy and stresses in terms of invariants of shear and area deformation. We generalize an expression of the initial shear modulus, which is independent of the number of molecular orientations within the network and also derive a simplified version of the model. We apply the model and its simplified version to analyze micropipette aspiration computationally and analytically and explore the effect of local cytoskeletal density change. We also explore the discrepancies among shear modulus values measured using different experimental techniques reported in the literature. We find that the model exhibits hardening behavior and can explain many of these discrepancies. Moreover, we find that the distribution of natural lengths plays a crucial role in the hardening behavior when the correct copy numbers of proteins are used. The initial shear modulus values we obtain using our current model (5.9–15.6 pN/μm) are close to the early estimates (6–9 pN/μm). This new, to our knowledge, constitutive model establishes a direct connection between the molecular structure of spectrin networks and constitutive laws and also defines a new picture of a much denser spectrin network than assumed in prior studies.

The Action of Cytomegalovirus (CMV) Promoter on Expression of Genetically Engineered Insulin in Rat Hepatocytes

Background: Insulin is a hormone produce in the pancreas that stimulates glucose uptake from blood to enter the body's cells, where it is converted into energy needed by muscles and tissues to function. The pancreatic cells are responsible for the secretion of insulin. In diabetes, the body cannot produce enough insulin or cannot use insulin effectively.
 Objectives: In this study, we attempted to activate the hepatic cells to secrete insulin instead of the pancreatic cells.
 Methodology: This was done by gene therapy; an excellent strategy to treat diabetes by supplying the correct wild type copy of a furin cleavable sites preproinsulin. The preproinsulin was extracted from the rat DNA, cloned and mutated to generate the two furin cleavable sites responsible for removal of C-peptide to form the two chains (A and B) for mature insulin production. This mutated insulin was derived by Cytomegalovirus (CMV) promoter to express insulin by its transfection inside the primary rat hepatocytes using a non-viral vehicle to keep the hepatic cells healthier against the transfection. In vitro, the rat hepatocytes could not divide well as in vivo, but with special hormones like insulin and dexamethasone lived longer and kept their function.
 Results: The CMV promoter is strong and lead to over expression of mature insulin inside rat hepatocytes leading to deterioration, toxicity of hepatocytes and finally cell death. Hepatocytes in vitro were more fragile and needed some modification to adapt with secretion of insulin.
 Conclusion: Glucokinase or glucose transporter promoters were much more perfect than CMV. They can activate the hepatocytes to modulate the glucose level and so limiting the amount of insulin secreted. These promoters are weaker than CMV but much more perfect for hepatocytes.

An Efficient Double-Layer Blockchain Method for Vaccine Production Supervision.

A vaccine is a biological product which is an important means for human beings to protect themselves. Most of its users are young children with weak immunity. Once a vaccine has a problem, it will pose a serious threat to the lives of many people. At present, the supervision of vaccine production is very simple. The vaccine production record is completely controlled by the enterprises. Enterprises only submit production records to the supervisory agency for review when the vaccine needs to be sold. Production records are easily forged and modified. In order to solve the shortcomings of traditional centralized management. We propose a supervision method for vaccine production based on double-level blockchain. At first, we have designed a double-level blockchain structure. The first level is private data of vaccine prduction enterprise, including production records and corresponding hash. The next level is public data, including production records hash and vaccine information. In this way, we make vaccine enterprise to submit production records in a timely manner without fear of privacy leaks. We avoid enterprise tampering or falsification of production records through the non-tampering features and time stamps of the blockchain. To improve the time efficiency, we propose a consensus mechanism for multi-node cooperate. The primary supervisory node provides sorting services and verifies the correctness of the blockchain replica. The ordinary supervisory node can replace the primary supervisory node when necessary, and help the primary supervisory node recovers data in case of information loss. The review node is responsible for providing complete and correct blockchain copies for other nodes. So we can avoids the problem of waste of time resources in the traditional blockchain system. In addition, in order to avoid the waste of space caused by the redundancy of the blockchain, we propose a vaccine data cutting mechanism. We use the timestamp of the blockchain and the vaccine validity period to determine if the block can be cutted. At the same time, it is also possible to judge whether the block can be cutted based on the information exchange with the vaccination institution. Through these methods, we have realized spatiotemporal efficiency supervision of vaccine production. And for the time being, research work in the field of vaccine production supervision is still very rare. So Our work is ground-breaking.

A Combined Technique of SC + MPC + APC to Achieve Higher Error Correction Probability and Throughput Over APC and MPC Techniques in a Wireless Network

In this paper, we present the combined techniques of Selection Combining (SC), Modified Packet Combining (MPC) and Aggressive Packet Combining (APC) for higher probability of packet error correction and packet throughput in three branches diversity system (multi-path) over conventional MPC and APC techniques. The three copies of the same packet are used to transmit from transmitter (Tx) to receiver (Rx) in three paths. Like in conventional SC technique, the Rx checks all the received copies individually from three paths whether the received copies are correct or not using frame checking sequences. If any one of the received copies is obtained, the correct copy then it will discard the remaining copies and request for next packet; otherwise the received erroneous copies will be combined. The combined erroneous copies perform ‘r’ number of pair-wise XORed after the erroneous copies are performed bit converted in MPC technique. If ‘r’ pair-wise XOR operations fail to recover the original copy, then APC technique implements with the converted bit stream. Thus, the simulation results using MATLAB obtain better performance in terms of probability of packet error correction and packet throughput over conventional MPC and APC schemes.

Correction for Kennan et al., "Genomic Evidence for a Globally Distributed, Bimodal Population in the Ovine Footrot Pathogen Dichelobacter nodosus".

Volume 5, issue 5, e01821-14, 2014, [https://doi.org/10.1128/mBio.01821-14][1]. It has been drawn to our attention that there are errors in the list of SRA accession numbers in Table S6 of this article, specifically in the last column of this table. Accordingly, we have provided a corrected copy of

British Society for Gene and Cell Therapy Autumn Conference Friday 23 November 2018 Regent's University Conference Centre, London, UK www.bsgct.org

British Society for Gene and Cell Therapy Autumn Conference Friday 23 November 2018 Regent's University Conference Centre, London, UK www.bsgct.org

Quantum Image Steganography Protocol Based on Quantum Image Expansion and Grover Search Algorithm

Quantum image steganography is one of the important research branches of quantum secure communications. In this paper, a large payload quantum image steganography protocol based on quantum image expansion and the Grover search algorithm is proposed. The new algorithm adopts quantum log-polar image (QUALPI) representation to prepare the quantum image before introducing a quantum expansion technique to form the superposition of multiple image copies with the same size angle difference as the carrier. Then, it embeds a secret message into one quantum image copy with a specific rotation angle encoded. In order to accurately extract the secret message embedded, the Grover search algorithm is used to locate the correct quantum image copy. Based on the quantum uncertainty and quantum non-cloning theorems, the new algorithm can not only achieve good imperceptibility and security but also large payload due to the algorithm's good coding scalability. The experimental results by performing MATLAB simulation prove the conclusions.

Novel gene based therapies for inherited retinal dystrophies in pediatric patients

SummaryGene based therapies for inherited retinal dystrophies have improved dramatically over the past 10 years. Clinical trials in patients based on the adeno‐associated virus (AAV) mediated gene transfer of a correct cDNA copy of the mutant gene were started for several disorders, including early onset severe retinal dystrophy associated with mutations in the RPE65 gene, and juvenile retinoschisis due to mutations in the RS1 gene. Furthermore, optogenetic approaches and cell based therapies are on the verge to the clinical application. The highest expectations though have raised the recent development of genome editing strategies based on CRISPR‐Cas endonucleases, which together with a template DNA enable the correction of the mutant DNA sequence ex vivo or in vivo in photoreceptor cells. In this lecture, an overview will be given on the current state of the art in gene based therapies for pediatric patients, and limitations and potential solutions will be discussed.

On the Phenomenology and Typology of Errors in Old Russian Apostolos Manuscripts from the 12th–14th Centuries

The article analyzes errors in two Old-Russian Apostolos manuscripts: Apostolus Christinopolitanus from the 12th century (an example of the commented type) and Tolstovskiy Apostolus from the 14th century (an example of the continuous type). The result of this research is new information about the reception of loan words, text reinterpretation, the influence of antigraphs and comments, and the personality of the Tolstovskiy scribe. The author corrects errors on both lexical and grammatical levels. Some of the lexical errors are provoked by deformations in the Greek text (homonym and paronym mixing). There are also various transformations of appellatives to onyms, and vice versa. All of this might occur in the antigraph as well as in the Tolstovskiy manuscript. Grammatical errors are fixed on the macro- and microlevels. There are cases of incorrect text segmentation within syntagms and between paragraphs in the Tolstovskiy manuscript which take place owing to the influence of the Praxapostolos and the commented type of Apostolos. Both manuscripts are also influenced by comments. This is reflected in grammatical and semantic alteration of the main text and, more rarely, in direct comments included in the main text, although the latter is not supposed for the continuous type to which the Tolstovskiy manuscript belongs. As for the chirographer of the Tolstovskiy manuscript, his background could not be characterized by a high level of literacy or by rich knowledge of the New Testament. He obviously did not have any Greek copies or more correct Slavonic copies at his disposal.

Surely, We’ll Need Backups

Abstract For 20 years the U. S. Newspaper and Brittle Books Programs focused national resources on a singular approach to preserving acidic paper in research libraries—converting printed “information” to microfilm. In many cases, the original historical material was discarded in the process. This method failed to address the root problem that could have been solved more effectively and more economically through preventive conservation. Such conservation was ignored because an ulterior motive to the brittle books “crisis” was to establish a national digital library. A serious consequence arising from this subterfuge is that today mid-sized research libraries are not committed to—and perhaps fail to comprehend the importance of—preserving their general collections. Collectively these diverse, paper-based collections represent a huge swath of the nation’s historical record. Falsely believing digital media will make paper obsolete, and motivated by a need to create space in libraries, today’s vigorous weeding for non-rare, paper-based general collections threatens to destroy evidence that will be required for research and future digitization projects. These materials will be needed as backups to accurately regenerate screen copies; as master copies to augment, enhance, or correct faulty screen copies; and, as authentication to verify original production techniques and determine provenance. Without careful coordination and item-level verification, today’s isolated weeding practices could eviscerate American general collections with devastating results for future scholarship, repeating the mistakes of the U. S. Newspaper and Brittle Books Programs.

Packet Combining and APC using Decode and Forward Relay over Wireless Networks

Decode and Forward (DF) is a type of opportunistic relay in which the packet is relayed only when the direct transmission of information from source to the destination fails. This results in bandwidth optimization of the overall system as well as considerable saving of resources by avoiding the unnecessary relay of information when it is not required. In this paper the authors have proposed a system using DF relay. By applying proper error correction techniques at the receiver side, it has been found that a corrected copy can be recovered from any erroneous receptions. Also the system offers much higher throughput and packet error rate (PER) performance, which is evident from the mathematical and simulation analysis made in the later part of this paper.

ZFN-Mediated Gene Targeting at the Albumin Locus in Liver Results in Therapeutic Levels of Human FIX in Mice and Non-Human Primates

Hemophilia is an attractive target for gene therapy, since activity levels as low as 1% to 2% of normal are beneficial and levels of ~5% prevent spontaneous bleeding. Our goal was to provide a single treatment that permanently enables hepatic production of therapeutic levels of hFIX activity to decrease or potentially eliminate the need for prophylactic treatment in hemophilia B patients. We performed targeted in vivo genome editing using 1) two zinc finger nucleases (ZFNs) targeting intron 1 of the albumin locus, and 2) a human F9 donor template construct. The ZFNs and donor template are encoded on separate hepatotropic adeno-associated virus serotype 2/6 (AAV2/6) vectors injected intravenously, resulting in targeted insertion of a corrected copy of the hF9 gene into the albumin locus in a proportion of liver hepatocytes. The albumin locus was selected as a "safe harbor" as production of this most abundant plasma protein exceeds 10 g/day, and moderate reductions in those levels are well-tolerated. These genome edited hepatocytes produce normal hFIX in therapeutic quantities, rather than albumin, driven by the highly active albumin enhancer/promoter, to treat hemophilia B; the genetic modification is expected to be sustained even in the face of hepatocyte turnover, making this approach attractive for treating young children with hemophilia before the appearance of significant organ damage.Transformed and primary human hepatocytes transduced in vitro with AAV2/6 encoding human albumin ZFNs and a promoterless hF9 transgene were shown to secrete hFIX. Extensive molecular analyses demonstrated that this was due to targeted integration of the hF9 transgene at the albumin locus and splicing of this gene into the albumin transcript. By employing AAV2/6 delivery of murine-specific ZFNs in vivo, stable levels of hFIX were observed in blood of mice injected with the albumin ZFNs and hF9 transgene donor. C57BL/6 mice were administered vehicle (n=20) or AAV2/6 vectors (n=25) encoding mouse surrogate reagents at 1.0 x1013 vector genome (vg)/kg via tail vein injection. ELISA analysis of plasma hFIX in the treated mice showed peak levels of 50-1053 ng/mL that were sustained for the duration of the 6-month study. Analysis of FIX activity from mouse plasma confirmed bioactivity commensurate with expression levels. Next, we report the feasibility of this approach in non-human primates (NHPs), showing that a single intravenous co-infusion of AAV2/6 vectors encoding the NHP targeted albumin-specific ZFNs and a human F9 donor at 1.2x1013 vg/kg (n=5/group) resulted in >50 ng/mL (>1% of normal) in this large animal model. The use of higher AAV2/6 doses (up to 1.5x1014 vg/kg) yielded plasma hFIX levels up to 1000 ng/ml (or 20% of normal) in several animals and up to 2000 ng/ml (or 50% of normal) in a single animal, for the duration of the study (3 months). The treatment was well tolerated in mice and NHPs, with no significant toxicological findings related to AAV2/6 ZFN + donor treatment in either species at therapeutic doses. Together, these data support a clinical trial to determine if a single co-administration of ZFN and donor AAV vectors is sufficient to enable therapeutic and potentially lifelong production of the clotting factor for the treatment of Hemophilia B. DisclosuresWechsler:Sangamo BioSciences: Employment. Meyer:Sangamo Biosciences Inc: Employment. Spratt:Sangamo Biosciences Inc: Employment. Greengard:Sangamo Biosciences Inc: Employment. Santiago:Sangamo Biosciences Inc: Employment. Sproul:Sangamo Biosciences Inc: Employment. Surosky:Sangamo Biosciences Inc: Employment. Paschon:Sangamo Biosciences Inc: Employment. Dubois-Stringfellow:Sangamo Biosciences Inc: Employment. Ando:Sangamo Biosciences Inc: Employment. Nichol:Sangamo Biosciences Inc: Employment. Rebar:Sangamo BioSciences: Employment. Holmes:Sangamo BioSciences: Employment.

Changing Biosynthetic Profiles by Expressing bldA in Streptomyces Strains.

Recently we described an unusual way of activating a cryptic gene cluster when we explored the origin of the bald phenotype of Streptomyces calvus. Complementation of S. calvus with a correct copy of bldA restored sporulation and additionally promoted production of a new natural products. In this study we report on the expression of bldA in several Streptomyces strains that have been described as "poorly sporulating" strains. In seven out of 15 cases, HPLC profiling revealed the production of new compounds, and in two cases the overproduction of known compounds. Two compounds were isolated and their structures were determined.

Novel Genome-Editing Tools to Model and Correct Primary Immunodeficiencies.

Severe combined immunodeficiency (SCID) and other severe non-SCID primary immunodeficiencies (non-SCID PID) can be treated by allogeneic hematopoietic stem cell (HSC) transplantation, but when histocompatibility leukocyte antigen-matched donors are lacking, this can be a high-risk procedure. Correcting the patient’s own HSCs with gene therapy offers an attractive alternative. Gene therapies currently being used in clinical settings insert a functional copy of the entire gene by means of a viral vector. With this treatment, severe complications may result due to integration within oncogenes. A promising alternative is the use of endonucleases such as ZFNs, TALENs, and CRISPR/Cas9 to introduce a double-stranded break in the DNA and thus induce homology-directed repair. With these genome-editing tools a correct copy can be inserted in a precisely targeted “safe harbor.” They can also be used to correct pathogenic mutations in situ and to develop cellular or animal models needed to study the pathogenic effects of specific genetic defects found in immunodeficient patients. This review discusses the advantages and disadvantages of these endonucleases in gene correction and modeling with an emphasis on CRISPR/Cas9, which offers the most promise due to its efficacy and versatility.