scholarly journals Gene editing in tree and clonal crops: progress and challenges

2021 ◽  
Author(s):  
Greg S. Goralogia ◽  
Thomas P. Redick ◽  
Steven H. Strauss
Keyword(s):  
Gene Editing ◽  
High Efficiency ◽  
National Environmental Policy Act ◽  
Functional Protein ◽  
Major Step ◽  
Insertion Event ◽  
Narrow Host Range ◽  
The Usa ◽  
Legal Innovation ◽  
Science And Risk

AbstractBecause of the limitations inherent in conventional breeding of trees and clonally propagated crops, gene editing is of great interest. Dozens of published papers attest to the high efficiency of CRISPR-based systems in clonal crops and trees. The opportunity for “clean” edits is expected to avoid or reduce regulatory burdens in many countries and may improve market acceptance. To date, however, nearly all studies in trees and clonal crops retained all of the gene editing machinery in the genome. Despite high gene editing efficiency, technical and regulatory obstacles are likely to greatly limit progress toward commercial use. Technical obstacles include difficult and slow transformation and regeneration, delayed onset of flowering or clonal systems that make sexual segregation of CRISPR-associated genes difficult, inefficient excision systems to enable removal of functional (protein- or RNA-encoding) transgenic DNA, and narrow host range or limited gene-payload viral systems for efficient transient editing. Regulatory obstacles include those such as in the EU where gene-edited plants are regulated like GMO crops, and the many forms of method-based systems that regulate stringently based on the method vs. product novelty and thus are largely applied to each insertion event. Other major obstacles include the provisions of the Cartagena Protocol with respect to international trade and the need for compliance with the National Environmental Policy Act in the USA. The USDA SECURE act has taken a major step toward a more science- and risk-based—vs. method and insertion event based—system, but much further regulatory and legal innovation is needed in the USA and beyond.

scholarly journals In vivo genome editing in mouse restores dystrophin expression in Duchenne muscular dystrophy patient muscle fibers

2021 ◽  
Vol 13 (1) ◽  
Author(s):  
Menglong Chen ◽  
Hui Shi ◽  
Shixue Gou ◽  
Xiaomin Wang ◽  
Lei Li ◽  
...  
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Mouse Model ◽  
Genome Editing ◽  
Large Scale ◽  
Gene Editing ◽  
High Efficiency ◽  
Muscle Fibers ◽  
Muscle Cells

Abstract Background Mutations in the DMD gene encoding dystrophin—a critical structural element in muscle cells—cause Duchenne muscular dystrophy (DMD), which is the most common fatal genetic disease. Clustered regularly interspaced short palindromic repeat (CRISPR)-mediated gene editing is a promising strategy for permanently curing DMD. Methods In this study, we developed a novel strategy for reframing DMD mutations via CRISPR-mediated large-scale excision of exons 46–54. We compared this approach with other DMD rescue strategies by using DMD patient-derived primary muscle-derived stem cells (DMD-MDSCs). Furthermore, a patient-derived xenograft (PDX) DMD mouse model was established by transplanting DMD-MDSCs into immunodeficient mice. CRISPR gene editing components were intramuscularly delivered into the mouse model by adeno-associated virus vectors. Results Results demonstrated that the large-scale excision of mutant DMD exons showed high efficiency in restoring dystrophin protein expression. We also confirmed that CRISPR from Prevotella and Francisella 1(Cas12a)-mediated genome editing could correct DMD mutation with the same efficiency as CRISPR-associated protein 9 (Cas9). In addition, more than 10% human DMD muscle fibers expressed dystrophin in the PDX DMD mouse model after treated by the large-scale excision strategies. The restored dystrophin in vivo was functional as demonstrated by the expression of the dystrophin glycoprotein complex member β-dystroglycan. Conclusions We demonstrated that the clinically relevant CRISPR/Cas9 could restore dystrophin in human muscle cells in vivo in the PDX DMD mouse model. This study demonstrated an approach for the application of gene therapy to other genetic diseases.

scholarly journals USA Gets Tough on Whales

Oryx ◽  
1971 ◽  
Vol 11 (2-3) ◽  
pp. 102-102
Author(s):  
Richard Fitter
Keyword(s):  
Blue Whale ◽  
Major Step ◽  
The Usa

The FPS Hon. Secretary reports on the International Whaling Commission's meeting in Washington which he attended on behalf of the Society. A major step forward was the decision to abolish the blue whale unit, but the quota fixed for the next season was again unrealistically high and the USA shows signs of losing patience.

scholarly journals Highly efficient generation of canker resistant sweet orange enabled by an improved CRISPR/Cas9 system

2021 ◽  
Author(s):  
Xiaoen Huang ◽  
Nian Wang
Keyword(s):  
Genome Editing ◽  
Gene Editing ◽  
Target Genes ◽  
High Efficiency ◽  
Sweet Orange ◽  
Susceptibility Gene ◽  
Transformation Rate ◽  
Biallelic Mutation ◽  
Important Species ◽  
Citrus Production

Sweet orange (Citrus sinensis) is the most economically important species for the citrus industry. However, it is susceptible to many diseases including citrus bacterial canker caused by Xanthomonas citri subsp. citri (Xcc) that triggers devastating effects on citrus production. Conventional breeding has not met the challenge to improve disease resistance of sweet orange due to the long juvenility and other limitations. CRISPR-mediated genome editing has shown promising potentials for genetic improvements of plants. Generation of biallelic/homozygous mutants remains difficult for sweet orange due to low transformation rate, existence of heterozygous alleles for target genes and low biallelic editing efficacy using the CRISPR technology. Here, we report improvements in the CRISPR/Cas9 system for citrus gene editing. Based on the improvements we made previously (dicot codon optimized Cas9, tRNA for multiplexing, a modified sgRNA scaffold with high efficiency, CsU6 to drive sgRNA expression), we further improved our CRISPR/Cas9 system by choosing superior promoters (CmYLCV or CsUbi promoter) to drive Cas9 and optimizing culture temperature. This system was able to generate a biallelic mutation rate of up to 89% for Carrizo citrange and 79% for Hamlin sweet orange. Consequently, this system was used to generate canker resistant Hamlin sweet orange by mutating the effector binding element (EBE) of canker susceptibility gene CsLOB1, which is required for causing canker symptoms by Xcc. Six biallelic Hamlin sweet orange mutant lines in the EBE were generated. The biallelic mutants are resistant to Xcc. Biallelic mutation of the EBE region abolishes the induction of CsLOB1 by Xcc. This study represents a significant improvement in sweet orange gene editing efficacy and generating disease resistant varieties via CRISPR-mediated genome editing. This improvement in citrus genome editing makes genetic studies and manipulations of sweet orange more feasible.

Design of Hydrogen Fuel Cell Autorickshaw

2006 ◽  
Author(s):  
Ikhlaq Khattak ◽  
Mirza Jamil Yousaf
Keyword(s):  
Fuel Cell ◽  
Air Quality ◽  
Internal Combustion Engines ◽  
Motor Vehicles ◽  
Oil Well ◽  
Hydrogen Fuel Cell ◽  
Major Step ◽  
Oil Mixtures ◽  
The Usa ◽  
Cleaner Technology

In Asia there are less private cars, but there is a high proportion of 2-stroke engines in scooters, motorcycles, auto-rickshaws (Tuk-Tuks), all running on petrol-oil mixtures with levels of hydrocarbon emissions (from partially burnt fuel and oil) well in excess of levels permitted in the USA and Europe. Worldwide Rickshaw/scooter/motorcycle type engine production is estimated at 17 million per year. According to National Transport Research Center (NTRC), the total population of registered (all types) motor vehicles in Pakistan in year 2000 was 4.224 million, out of which more than half of the population is (2.206 million) two wheelers or three wheelers (motorcycle/scooter/auto rickshaw). Almost all auto rickshaws have two stroke power packs and also 60% of motorcycle/scooters are of the same category. Pakistan is a very densely populated developing country, with very loose environment protection rules, which are practically unregulated due to large financial implications. This scenario leads to adverse air quality conditions especially in large cities of the country where the main contributory factors are vehicular traffic, that too, two stroke vehicles Industry, diesel-powered vehicles, and the omnipresent three-wheeled, two-stroke rickshaws all contribute to the extremely dirty air. Taxi/car use is increasing, but rickshaws have the advantage of being able to swarm through the congested car traffic in cities. This explains the over .6 million motorcycles/scooters/rickshaws currently in Pakistan, of which approximately 20% are two stroke Auto-rickshaws of 175 cc. Pakistan’s vehicle fleet has a growth rate of 8.0% (1990–99). The purpose of this study is to examine a particular application of fuel cell technology “The Auto Rickshaws”. They are small three-wheeled vehicles that can carry three people. Due to their small size and low price, rickshaws have traditionally been powered by high power density two-stroke internal combustion engines. Two-stroke engines produce a great deal of pollution and are an object of concern in many Asian countries. Severe pollution from two-stroke engines is a significant driver for cleaner technology. Thus, the target of this study is the Asian urban commuter, since a rickshaw is largely used in many Asian cities and contributes directly to air pollution in major crowded cities of Pakistan also. Countries like China, India, Bangladesh, Taiwan and Pakistan [1] are facing dramatic growth rates in two-stroke vehicle population as bicycle rickshaws are being replaced, so, low-powered but clean rickshaws would be a major step in providing mobility without compromising urban air quality.

scholarly journals Development of a CRISPR-SaCas9 system for projection- and function-specific gene editing in the rat brain

2020 ◽  
Vol 6 (12) ◽  
pp. eaay6687 ◽  
Author(s):  
Haojie Sun ◽  
Su Fu ◽  
Shuang Cui ◽  
Xiangsha Yin ◽  
Xiaoyan Sun ◽  
...  
Keyword(s):  
Rat Brain ◽  
Gene Editing ◽  
High Efficiency ◽  
Protein A ◽  
Cell Types ◽  
Cell Labeling ◽  
Specific Gene ◽  
Creb Binding Protein ◽  
A Genome ◽  
And Function

A genome editing technique based on the clustered regularly interspaced short palindromic repeats (CRISPR)–associated endonuclease Cas9 enables efficient modification of genes in various cell types, including neurons. However, neuronal ensembles even in the same brain region are not anatomically or functionally uniform but divide into distinct subpopulations. Such heterogeneity requires gene editing in specific neuronal populations. We developed a CRISPR-SaCas9 system–based technique, and its combined application with anterograde/retrograde AAV vectors and activity-dependent cell-labeling techniques achieved projection- and function-specific gene editing in the rat brain. As a proof-of-principle application, we knocked down the cbp (CREB-binding protein), a sample target gene, in specific neuronal subpopulations in the medial prefrontal cortex, and demonstrated the significance of the projection- and function-specific CRISPR-SaCas9 system in revealing neuronal and circuit basis of memory. The high efficiency and specificity of our projection- and function-specific CRISPR-SaCas9 system could be widely applied in neural circuitry studies.

scholarly journals Analysis of roof greening technology impact on rain and meltwater retention

2020 ◽  
Vol 175 ◽  
pp. 11023
Author(s):  
Elena Sysoeva ◽  
Margarita Gelmanova
Keyword(s):  
High Efficiency ◽  
Green Roof ◽  
Experimental Studies ◽  
Green Roofs ◽  
Climatic Conditions ◽  
Water Runoff ◽  
Substrate Thickness ◽  
Runoff Reduction ◽  
The Usa ◽  
Analytical Review

Over the past 20 years, a large number of studies have been published on reducing storm runoff by various types of green roofs. This article analyzes the results of experimental studies presented in 39 publications on green roof runoff reduction in a climate similar to the climate of Russia: in Canada, the USA, Finland, Norway, France. An analytical review found that the ability of green roofs to retain rainfall varies from 20 to 99.5% depending on climatic conditions (duration and intensity of rains, duration of dry periods, solar radiation, temperature and humidity, wind conditions), the properties of green roof layers (moisture capacity of the substrate and a drainage layer, the substrate thickness), the type of vegetation, the geometry of a green roof (slope and orientation). Green roofs can be a useful tool for reducing urban storm water runoff. However, in order to ensure high efficiency, it is necessary to use green roof technology with other measures to reduce runoff.

scholarly journals Simplified pipelines for genetic engineering of mammalian embryos by CRISPR-Cas9 electroporation†

2019 ◽  
Vol 101 (1) ◽  
pp. 177-187 ◽  
Author(s):  
Deqiang Miao ◽  
Mariana Ianello Giassetti ◽  
Michela Ciccarelli ◽  
Blanca Lopez-Biladeau ◽  
Jon M Oatley
Keyword(s):  
Embryo Transfer ◽  
Gene Editing ◽  
High Efficiency ◽  
Mammalian Species ◽  
Culture Conditions ◽  
Surgical Approaches ◽  
Production Traits ◽  
Enhanced Production ◽  
Mammalian Embryos ◽  
Transfer Method

Abstract Gene editing technologies, such as CRISPR-Cas9, have important applications in mammalian embryos for generating novel animal models in biomedical research and lines of livestock with enhanced production traits. However, the lack of methods for efficient introduction of gene editing reagents into zygotes of various species and the need for surgical embryo transfer in mice have been technical barriers of widespread use. Here, we described methodologies that overcome these limitations for embryos of mice, cattle, and pigs. Using mutation of the Nanos2 gene as a readout, we refined electroporation parameters with preassembled sgRNA-Cas9 RNPs for zygotes of all three species without the need for zona pellucida dissolution that led to high-efficiency INDEL edits. In addition, we optimized culture conditions to support maturation from zygote to the multicellular stage for all three species that generates embryos ready for transfer to produce gene-edited animals. Moreover, for mice, we devised a nonsurgical embryo transfer method that yields offspring at an efficiency comparable to conventional surgical approaches. Collectively, outcomes of these studies provide simplified pipelines for CRISPR-Cas9-based gene editing that are applicable in a variety of mammalian species.

scholarly journals Ethical and Welfare Implications of Genetically Altered Non-Human Primates for Biomedical Research

2020 ◽  
pp. 1-26
Author(s):  
Mark J. Prescott
Keyword(s):  
Biomedical Research ◽  
Gene Editing ◽  
Ethical Issues ◽  
Neurological Diseases ◽  
Therapeutic Interventions ◽  
Large Numbers ◽  
The Usa ◽  
Welfare Implications ◽  
Models Of Disease ◽  
Human Primate

Abstract Breakthroughs in gene editing technologies have made it feasible to create genetically altered (GA) non-human primate (NHP) models of disease. This area of research is accelerating, particularly in China, Japan and the USA, and could lead to an increase in NHP use globally. The hope is that genetic models in animal species closely related to humans will significantly improve understanding of neurological diseases and validation of potential therapeutic interventions, for which there is a dire need. However, the creation and use of GA NHPs raises serious animal welfare and ethical issues, which are highlighted here. It represents a step change in how these highly sentient animals are used in biomedical research, because of the large numbers required, inherent wastage and the sum of the harms caused to the animals involved. There is little evidence of these important issues being addressed alongside the rapidly advancing science. We are still learning about how gene editing tools work in NHPs, and significant added scientific and medical benefit from GA NHP models has yet to be demonstrated. Together, this suggests that current regulatory and review frameworks, in some jurisdictions at least, are not adequately equipped to deal with this emerging, complex area of NHP use.

scholarly journals High-efficiency CRISPR/Cas9 multiplex gene editing using the glycine tRNA-processing system-based strategy in maize

2016 ◽  
Vol 16 (1) ◽  
Author(s):  
Weiwei Qi ◽  
Tong Zhu ◽  
Zhongrui Tian ◽  
Chaobin Li ◽  
Wei Zhang ◽  
...  
Keyword(s):  
Gene Editing ◽  
High Efficiency ◽  
Processing System ◽  
Trna Processing

Detection of Radon-Progeny and Other Alpha-Emitting Radionuclides in Air Using Tensioned Metastable Fluid Detectors

2017 ◽  
Author(s):  
N. Boyle ◽  
B. Archambault ◽  
A. Hagen ◽  
C. Meert ◽  
R. P. Taleyarkhan
Keyword(s):  
Environmental Protection Agency ◽  
Temperature Effects ◽  
High Efficiency ◽  
Collection Efficiency ◽  
Nuclear Industry ◽  
Reference Laboratory ◽  
Neutron Fission ◽  
Relative Standard ◽  
Gas Seepage ◽  
The Usa

Alpha radiation emitting radon (Rn) gas seepage into homes in the USA leads to over 21,000 annual lung cancer deaths (according to the US-Environmental Protection Agency, EPA) leading to mandatory monitoring for Rn throughout the USA. In the nuclear industry alpha emitting radionuclides in air (e.g., in spent fuel reprocessing) also constitute a major safety and security-safeguards related issues. Purdue University, along with Sagamore Adams Laboratories LLC, is developing the tensioned metastable fluid detector (TMFD) technology for general-purpose alpha-neutron-fission spectroscopy. This paper focuses on rapid, high-efficiency detection of Rn and progeny in air using the novel TMFD technology; Rn and progeny isotopes in air are sparged through the TMFD detection fluid (to entrap the radioactive gas), which is then placed under a metastable state. Through tailoring the metastable fluid state, an audible and visible cavitation detection event is created and readily detected from transient bubble formation. Changing the tensioned state allows for the spectroscopic differentiability of Rn and its daughters which can be used to actively measure the equilibrium between the parent and daughter products. Such a technique can also be used to monitor the atmosphere in critical nuclear facilities for contamination from other alpha emitting isotopes (e.g., Pu, Cm, U...). TMFDs offer the unique ability for high intrinsic efficiency (>95%) alpha-neutron-fission fragment detection, while remaining blind to background beta and gamma radiation (qualified to >3.8×108 Bq m−3 using a dissolved 32P beta source, and also via gammas from a 53 R/hr 137Cs gamma source). Immunity to beta and gamma is beneficial for the discrimination of buildup of beta-emitting Thoron and Rn progeny in the detection fluid allowing for reusability. This paper will discuss the research results pertaining to detection of Radon and progeny in air, for concentrations between 74 Bq m−3 (2 pCi/L) and 740 Bq m−3 (20 pCi/L). The system measures a radon concentration between these levels to within ±15% intrinsic relative error (IRE) within 24 hours meeting the standards outlined by the American Association of Radon Scientists and Technicians-National Radon Proficiency Program (AARST-NRPP) Device Evaluation Program. Precision evaluation was also performed and the relative standard deviation defined by the AARST-NRPP was <5% exceeded the requirement of 25%. Ambient temperature effects were assessed at 10 °C and at 27 °C, which revealed a large increase in collection efficiency with decreasing sampling temperature and slight increase with increasing sampling temperature. Temperature effects on sensitivity thresholds and volumetric expansion were measured and used to compensate for variability in temperature over time. Blind testing with the help of Bowser-Morner Radon Reference Laboratory was performed and succeeded in accurately determining the Rn in air concentration to within 20% within only 6h of sampling. Finally, a 48-hour based collection time has also been developed for use in dwellings where Rn in air concentrations may vary in a day. Overall, the reproducibility and precision of TMFD technology is found to allow for an efficient, cost-effective, reliable, and environmentally friendly means of Rn and progeny detection, and by extension for use for general actinide in air monitoring for the nuclear industry.

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