Gene and genome editing最新文献

Genome editing is a widely used tool for making precise genomic changes. However, no specialized databases which are sufficiently comprehensive are available with consolidated data on genome editing. Therefore, we have developed a genome editing meta-database (GEM, web interface: https://bonohu.hiroshima-u.ac.jp/gem/) that aims to collect an exhaustive dataset of metadata related to genome editing. The GEM is systematically extracted via specialized text-mining from PubMed and PubMed Central (PMC) literatures that contain experiments involving the use of the seven types of genome editing tools. We constructed a dataset consisting of 50,162 entries of metadata based on 15,952 studies with 1,088 species that users can search and retrieve graphically in the GEM web interface. GEM, therefore, allows bioscientists to easily obtain information about genome editing and apply it to their respective research.

RNA molecules regulate and participate in a vast array of cellular processes, and the scientific community is now well into a new era in which some aspect of RNA biology—as a tool, therapeutic, diagnostic, or central player in fundamental biological processes—is becoming increasingly important. Any abnormality in RNA often results in a deficiency of protein production, which may also cause various diseases. Among the various types of RNA processing, RNA editing is an enigmatic reaction in which the sequence context at the RNA level is rewritten. This review summarizes our current understanding of RNA editing in various organisms especially focusing on C-to-U and U-to-C RNA editing in plants and pentatricopeptide repeat (PPR) proteins that are responsible for target RNA recognition and editing reactions. An overview of the recent developments in synthetic RNA editing tools and future perspective of the use of PPR system for gene therapy is also provided.

Deep sequencing technology in forward genetics is a powerful tool to identify causal mutations underlying hereditary human diseases. To elucidate the etiological mechanisms, reverse genetics in human cultured cells is useful for generating disease models in vitro. However, the development of reverse genetics has been slow because of the lower efficacy of homologous recombination in almost all mammalian cultured cells. The history of reverse genetics in cultured cells began with the advent of genome editing technology, which could effectively modify the genome via artificial nuclease-induced local DNA repair activity. Bidirectional genetics based on deep sequencing technology and genome editing technology is now an essential approach for clarifying the pathophysiology of hereditary diseases. Here, we provide an overview of the validity of genome editing in cultured cells and its technical problems, discussing the example of centrosome/cilia-related disease models in cultured cells.

The development of rapid, sensitive, specific and accurate diagnostic tests is essential for improving the treatment outcome of diseases. In the majority of disease diagnosis, nucleic acid-based tests are accepted as a gold standard. In general, these tests provide reliable results, yet they require highly trained personnel and specialized equipmentation. With the introduction of clustered regularly interspaced short palindromic repeats (CRISPR)-Cas systems in diagnostic applications, achieving breakthrough improvements in diagnostic tests is now possible. The specific target sequence recognition ability and trans-cleavage activity of certain Cas proteins enable novel applications of these systems in the development and improvement of diagnostic tests. These improvements and innovations allow for improved sensitivity, specificity and accuracy of point-of-care tests while keeping their costs at affordable levels. In this review, a comprehensive analysis of the common CRISPR-Cas systems used in diagnostic applications and the utilization of these systems in the design of novel biosensors is provided.

Genome editing technologies have brought dramatic changes in many fields of research, including plant sciences. Zinc finger nuclease, transcription activator-like effector nuclease (TALEN) and clustered regularly interspaced palindromic repeats (CRISPR)-CRISPR-associated protein 9 (Cas9) are key players in genome editing and have been developed for targeted mutagenesis. To apply genome editing to plants, optimization and development of several technologies to overcome plant-specific hurdles has been required. In this review, we highlight recent topics in the plant genome editing field in Japan, ranging from the development of a new genome editing tool to commercial applications of genome edited plants. Such achievements contribute greatly to the development of plant genome research and its application to plant breeding.

Gene knock-in can be defined as the introduction of precisely determined modifications, insertions, or replacements to the genome, which enables the generation of reporter cells, disease modeling and correction, humanization of animal cells and organisms, and so on. To date, gene knock-in systems have reached the fourth stage; i.e., the first, second, and third stages depend on unconstrained homologous recombination (HR)-mediated strategy, genome editing-assisted HR, and genome editing with various DNA double-strand break (DSB) repair pathways such as non-homologous end-joining and microhomology-mediated end-joining, respectively. Finally, in the fourth stage, DSB-free precision gene editors such as base editor and prime editor became available. These diversified strategies open up a new era of intentional editing of the genome, widely contributing to the functional genomics study.

Background

Innovation in gene therapy and genome editing raises high expectations for therapeutic breakthroughs. With the increasing maturity of the field, gene products using “gene transfer” technology, such as viral vectors, designer nucleases, incl. CRISPR/Cas, as the most recent, are frequently tested in clinical trials. Before such trials are launched, the anticipated risks and benefits of using gene transfer technologies must be evaluated to ascertain an ethical balance of risks and benefits.

Methods

We conducted semi-structured interviews with experts (n=15) in gene therapy/genome editing. We applied thematic text analysis to identify the qualitative spectrum of strengths, weaknesses, opportunities, and threats (SWOT) of a risk assessment approach to gene therapy/genome editing research based on a comprehensive set of nine mechanistic categories of adverse reactions combined with estimates of risk probability according to World Health Organization (WHO) adverse reaction terminology.

Results

Our study revealed a clear demand for a structured approach to risk assessment gene therapy/genome editing. The interviews indicate that the nine presented mechanistic categories may be helpful to structure this risk assessment prior to initiating a new study. The interviews revealed a broad spectrum of practice-oriented SWOT, described in detail in this manuscript.

Discussion

The here presented SWOT for a structured approach to risk assessment prior to clinical trials with gene therapy/genome editing inform the refinement and implementation of such standardized approaches and the discussion among researchers, regulators, and funders. To overcome potential weaknesses and threats in the application of such a risk-based approach, the mechanistic categories need to be case-sensitive and complemented by information on the validity of relevant animal models, long-term risks, and information about patient characteristics.

Bioinformatics has become an indispensable technology in molecular biology for genome editing. In this review, we outline various bioinformatic techniques necessary for genome editing research. We first review state-of-the-art computational tools developed for genome editing studies. We then introduce a bio-digital transformation (BioDX) approach, which fully utilizes existing databases for biological innovation, and uses publicly available bibliographic full-text data and transcriptome data to survey genome editing target genes in model organism species, where substantial genomic information and annotation are readily available. We also discuss genome editing attempts in species with almost no genomic information. The transcriptome data, sequenced genomes, and functional annotations for these species are described, with a primary focus on the bioinformatic tools used for these analyses. Finally, we conclude on the need to maintain a database of genome editing resources for future development of genome editing research. Our review shows that the integration and maintenance of useful resources remains a challenge for bioinformatics research in genome editing, and that it is crucial for the research community to work together to create and maintain such databases in the future.

Epigenome editing is a technique by which the epigenome of a specific DNA may be manipulated. The application of this technology to mouse zygotes or mouse embryonic stem cells (mESC) to produce mice with modified epigenomes would allow determination of the phenotypic role of the epigenome of a specific region for a given gene. In other words, reverse epigenetics becomes possible. In this review, we will discuss the significance of epigenome editing, the methods available, and the application of epigenome editing in mice.

Chicken (Gallus gallus) is a major protein source and an important model organism in the avian species. Although genome editing has enabled genetic modifications of various organisms and has a significant impact on academia and industry, its application to chickens has been comparatively delayed owing to difficulties in handling their one-cell fertilized eggs. Thus, researchers have attempted to produce genome-edited chickens using primordial germ cells (PGCs), which are precursor cells of sperm or eggs. Currently, genome-edited chickens can be produced with the development of avian biotechnologies, PGCs culture methods, and germline chimerism systems, in particular. In this review, we describe the current status of genome editing in chickens, including avian biotechnologies, with a primary focus on the achievements of Japanese researchers. In addition, we discuss the remaining issues and make suggestions for future research.