scholarly journals Transcription activator–like effector nuclease (TALEN)-mediated genome editing to generate genetically modified monkeys

2014 ◽  
Vol 7 (10) ◽  
pp. 296-296
Keyword(s):  
Genome Editing ◽  
Genetically Modified ◽  
Transcription Activator

31 PRODUCTION EFFICIENCY OF GENE KNOCKOUT PIGS USING GENOME EDITING AND SOMATIC CELL CLONING

2015 ◽  
Vol 27 (1) ◽  
pp. 108
Author(s):  
H. Matsunari ◽  
M. Watanabe ◽  
K. Nakano ◽  
A. Uchikura ◽  
Y. Asano ◽  
...  
Keyword(s):  
Somatic Cell ◽  
Genome Editing ◽  
Production Efficiency ◽  
Gene Knockout ◽  
Genetically Modified ◽  
Zinc Finger Nucleases ◽  
International Institute ◽  
Induced Mutations ◽  
Transcription Activator

Genome editing technologies have been used as a powerful strategy for the generation of genetically modified pigs. We previously developed genetically modified clone pigs with organogenesis-disabled phenotypes, as well as pigs exhibiting diseases with similar features to those of humans. Here, we report the production efficiency of various gene knockout cloned pigs from somatic cells that were genetically modified using zinc finger nucleases (ZFN) or transcription activator-like effector nucleases (TALEN). The ZFN- or TALEN-encoding mRNAs, which targeted 7 autosomal or X-linked genes, were introduced into porcine fetal fibroblast cells using electroporation. Clonal cell populations carrying induced mutations were selected after limiting dilution. The targeted portion of the genes was amplified using PCR, followed by sequencing and mutation analysis. Among the collected knockout cell colonies, cells showing good proliferation and morphology were selected and used for somatic cell nuclear transfer (SCNT). In vitro-matured oocytes were obtained from porcine cumulus-oocyte complexes cultured in NCSU23-based medium and were used to obtain recipient oocytes for SCNT after enucleation. SCNT was performed as reported previously (Matsunari et al. 2008). The cloned embryos were cultured for 7 days in porcine zygote medium (PZM)-5 to assess their developmental ability. Cloned embryos were transplanted into the oviduct or uterus of oestrus-synchronized recipient gilts to evaluate their competence to develop to fetuses or piglets. Cloned embryos reconstructed with 7 types of knockout cells showed equal development to blastocysts compared with those derived from the wild-type cells (54.5–83.3% v. 60.7%). Our data (Table 1) demonstrated that the reconstructed embryos derived from knockout cells could efficiently give rise to cloned offspring regardless of the type of genome editing methodology (i.e. ZFN or TALEN). Table 1.Production efficiency of gene knockout cloned pigs using genome editing This study was supported by JST, ERATO, the Nakauchi Stem Cell and Organ Regeneration Project, JST, CREST, Meiji University International Institute for Bio-Resource Research (MUIIBR), and JSPS KAKENHI Grant Number 26870630.

scholarly journals Evolution and Application of Genome Editing Techniques for Achieving Food and Nutritional Security

2021 ◽  
Vol 22 (11) ◽  
pp. 5585
Author(s):  
Sajid Fiaz ◽  
Sunny Ahmar ◽  
Sajjad Saeed ◽  
Aamir Riaz ◽  
Freddy Mora-Poblete ◽  
...  
Keyword(s):  
Abiotic Stress ◽  
Food Security ◽  
Plant Breeding ◽  
Genome Editing ◽  
Crop Improvement ◽  
Hybrid Seed Production ◽  
Biotic And Abiotic Stress ◽  
Transcription Activator ◽  
Protein System ◽  
Breeding Techniques

A world with zero hunger is possible only through a sustainable increase in food production and distribution and the elimination of poverty. Scientific, logistical, and humanitarian approaches must be employed simultaneously to ensure food security, starting with farmers and breeders and extending to policy makers and governments. The current agricultural production system is facing the challenge of sustainably increasing grain quality and yield and enhancing resistance to biotic and abiotic stress under the intensifying pressure of climate change. Under present circumstances, conventional breeding techniques are not sufficient. Innovation in plant breeding is critical in managing agricultural challenges and achieving sustainable crop production. Novel plant breeding techniques, involving a series of developments from genome editing techniques to speed breeding and the integration of omics technology, offer relevant, versatile, cost-effective, and less time-consuming ways of achieving precision in plant breeding. Opportunities to edit agriculturally significant genes now exist as a result of new genome editing techniques. These range from random (physical and chemical mutagens) to non-random meganucleases (MegaN), zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), clustered regularly interspaced short palindromic repeats (CRISPR)/associated protein system 9 (CRISPR/Cas9), the CRISPR system from Prevotella and Francisella1 (Cpf1), base editing (BE), and prime editing (PE). Genome editing techniques that promote crop improvement through hybrid seed production, induced apomixis, and resistance to biotic and abiotic stress are prioritized when selecting for genetic gain in a restricted timeframe. The novel CRISPR-associated protein system 9 variants, namely BE and PE, can generate transgene-free plants with more frequency and are therefore being used for knocking out of genes of interest. We provide a comprehensive review of the evolution of genome editing technologies, especially the application of the third-generation genome editing technologies to achieve various plant breeding objectives within the regulatory regimes adopted by various countries. Future development and the optimization of forward and reverse genetics to achieve food security are evaluated.

scholarly journals Production of α1,3-galactosyltransferase targeted pigs using transcription activator-like effector nuclease-mediated genome editing technology

2016 ◽  
Vol 17 (1) ◽  
pp. 89 ◽  
Author(s):  
Jung-Taek Kang ◽  
Dae-Kee Kwon ◽  
A-Rum Park ◽  
Eun-Jin Lee ◽  
Yun-Jin Yun ◽  
...  
Keyword(s):  
Genome Editing ◽  
Transcription Activator

scholarly journals TALEN and CRISPR/Cas Genome Editing Systems: Tools of Discovery

Acta Naturae ◽  
2014 ◽  
Vol 6 (3) ◽  
pp. 19-40 ◽  
Author(s):  
A. A. Nemudryi ◽  
K. R. Valetdinova ◽  
S. P. Medvedev ◽  
S. M. Zakian
Keyword(s):  
Genome Editing ◽  
Dna Sequences ◽  
Genome Engineering ◽  
Disease Modeling ◽  
Regulation Of Gene Expression ◽  
Hereditary Disease ◽  
Transcription Activator ◽  
Wide Range ◽  
High Standards ◽  
Phenotype Formation

Precise studies of plant, animal and human genomes enable remarkable opportunities of obtained data application in biotechnology and medicine. However, knowing nucleotide sequences isnt enough for understanding of particular genomic elements functional relationship and their role in phenotype formation and disease pathogenesis. In post-genomic era methods allowing genomic DNA sequences manipulation, visualization and regulation of gene expression are rapidly evolving. Though, there are few methods, that meet high standards of efficiency, safety and accessibility for a wide range of researchers. In 2011 and 2013 novel methods of genome editing appeared - this are TALEN (Transcription Activator-Like Effector Nucleases) and CRISPR (Clustered Regulatory Interspaced Short Palindromic Repeats)/Cas9 systems. Although TALEN and CRISPR/Cas9 appeared recently, these systems have proved to be effective and reliable tools for genome engineering. Here we generally review application of these systems for genome editing in conventional model objects of current biology, functional genome screening, cell-based human hereditary disease modeling, epigenome studies and visualization of cellular processes. Additionally, we review general strategies for designing TALEN and CRISPR/Cas9 and analyzing their activity. We also discuss some obstacles researcher can face using these genome editing tools.

Concepts of Molecular Plant Breeding and Genome Editing

2021 ◽  
pp. 1-15
Keyword(s):  
Plant Breeding ◽  
Genome Editing ◽  
Genetic Linkage ◽  
Selection Process ◽  
Crop Improvement ◽  
Zinc Finger Nucleases ◽  
Induced Mutations ◽  
Transcription Activator ◽  
Targeted Gene Editing ◽  
Breeding Techniques

Traditional plant breeding depends on spontaneous and induced mutations available in the crop plants. Such mutations are rare and occur randomly. By contrast, molecular breeding and genome editing are advanced breeding techniques that can enhance the selection process and produce precisely targeted modifications in any crop. Identification of molecular markers, based on SSRs and SNPs, and the availability of high-throughput (HTP) genotyping platforms have accelerated the process of generating dense genetic linkage maps and thereby enhanced application of marker-assisted breeding for crop improvement. Advanced molecular biology techniques that facilitate precise, efficient, and targeted modifications at genomic loci are termed as “genome editing.” The genome editing tools include “zinc-finger nucleases (ZNFs),” “transcription activator-like effector nucleases (TALENs),” oligonucleotide-directed mutagenesis (ODM), and “clustered regularly interspersed short palindromic repeats (CRISPER/Cas) system,” which can be used for targeted gene editing. Concepts of molecular plant breeding and genome editing systems are presented in this chapter.

scholarly journals Progress and Challenges in the Improvement of Ornamental Plants by Genome Editing

Plants ◽  
2020 ◽  
Vol 9 (6) ◽  
pp. 687 ◽  
Author(s):  
Chang Ho Ahn ◽  
Mummadireddy Ramya ◽  
Hye Ryun An ◽  
Pil Man Park ◽  
Yae-Jin Kim ◽  
...  
Keyword(s):  
Genome Editing ◽  
Gene Editing ◽  
Genetically Modified Organisms ◽  
Ornamental Plants ◽  
Safety Evaluation ◽  
Evaluation Process ◽  
Vase Life ◽  
Transcription Activator ◽  
A Genome ◽  
Dna Specificity

Biotechnological approaches have been used to modify the floral color, size, and fragrance of ornamental plants, as well as to increase disease resistance and vase life. Together with the advancement of whole genome sequencing technologies, new plant breeding techniques have rapidly emerged in recent years. Compared to the early versions of gene editing tools, such as meganucleases (MNs), zinc fingers (ZFNs), and transcription activator-like effector nucleases (TALENs), clustered regularly interspaced short palindromic repeat (CRISPR) is capable of altering a genome more efficiently and with higher accuracy. Most recently, new CRISPR systems, including base editors and prime editors, confer reduced off-target activity with improved DNA specificity and an expanded targeting scope. However, there are still controversial issues worldwide for the recognition of genome-edited plants, including whether genome-edited plants are genetically modified organisms and require a safety evaluation process. In the current review, we briefly summarize the current progress in gene editing systems and also introduce successful/representative cases of the CRISPR system application for the improvement of ornamental plants with desirable traits. Furthermore, potential challenges and future prospects in the use of genome-editing tools for ornamental plants are also discussed.

31 EFFICIENT GENERATION OF klotho MUTATIONS IN PORCINE SOMATIC CELL NUCLEAR TRANSFER EMBRYOS USING A DELIVERY OF Cas9 RIBONUCLEOPROTEINS

2017 ◽  
Vol 29 (1) ◽  
pp. 123
Author(s):  
S. Lee ◽  
M. H. Jung ◽  
H. J. Oh ◽  
O.-J. Koo ◽  
B. C. Lee
Keyword(s):  
Somatic Cell ◽  
Genome Editing ◽  
Somatic Cell Nuclear Transfer ◽  
Nuclear Transfer ◽  
Premature Aging ◽  
Human Diseases ◽  
Limited Information ◽  
Transcription Activator ◽  
Klotho Gene ◽  
Donor Cells

Pigs are useful models for studying human diseases because of the similarity of their anatomy and physiology. Recent advances in genome editing techniques such as zinc-finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and clustered regularly interspaced short palindromic repeat-associated Cas9 system (CRISPR/Cas9) have made it possible to produce animals for specific purposes. Especially, recent application of the CRISPR/Cas9 system improved the efficiency of genome editing in pigs with higher targeting efficiency or percentage of desired mutation compared to other meganucleases (ZFNs and TALENs). The klotho deficiency in small animals such as mice is characterised by an extremely shortened life span with multiple aging-like phenotypes similar to human premature-aging syndromes. However, limited information is available on the function of klotho in large animals such as pigs. The objective of this study was to determine whether the use of non-selected porcine fibroblasts electroporated with Cas9/sgRNA ribonucleoproteins, targeting the klotho gene, for somatic cell nuclear transfer (SCNT) results in high mutation rates in embryos. A CRISPR sgRNA specific for the klotho gene was designed and sgRNA (targeting exon 3 of klotho) and type 2 Cas9 RNPs (total 36 μg, 1:4 ratio, respectively) were transfected into porcine fibroblasts via Neon (Life Technologies) with a single DC pulse of 1400 V for 30 ms. Then, transfected fibroblasts were cultured for 1 day and used randomly for SCNT without selection. SCNT was performed by enucleation of in vitro-matured porcine oocyte, followed by injection of non-selected donor cells, fusion with a single DC pulse of 200 V/mm for 30 μs using an electro cell fusion generator (LF101; Nepa Gene Co.), and electrical activation with a single DC pulse of 150 V/mm for 60 μs using a BTX Electro-Cell Manipulator 2001 (BTX Inc.). The SCNT embryos were cultured in PZM5 culture medium to Day 7 and analysed for the presence of modifications to the klotho gene. Blastocysts were classified as modified if they contained an INDEL as measured by both T7E1 assay and deep sequencing of PCR amplicons spanning the targeted exon. The klotho modification rate was 65% (n = 13), of which 38.5% (n = 5) of the embryos contained biallelic modifications. In conclusion, SCNT with non-selected donor cells transfected with Cas9/sgRNA RNPs might be an efficient and simple tool to produce klotho deficient pigs as models for human diseases. Further studies are required to generate klotho deficient pigs by performing embryo transfer to the recipients. This study was supported by Korea Institute of Planning and Evaluation for Technology in food, agriculture, forestry and fisheries (#311011–05–5-SB010, #114059–03–2-SB010), Research Institute for Veterinary Science, TS Corporation and the BK21 plus program.

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