scholarly journals Agroinfiltration Mediated Scalable Transient Gene Expression in Genome Edited Crop Plants

2021 ◽  
Vol 22 (19) ◽  
pp. 10882
Author(s):  
Maninder Kaur ◽  
Pooja Manchanda ◽  
Anu Kalia ◽  
Farah K. Ahmed ◽  
Eugenie Nepovimova ◽  
...  
Keyword(s):  
Genome Editing ◽  
Transient Gene Expression ◽  
Transformation Method ◽  
Pilot Scale ◽  
Dna Integration ◽  
Agrobacterium Mediated Transformation ◽  
Direct Delivery ◽  
Foreign Genes ◽  
Scale Experiment ◽  
Stable Genome

Agrobacterium-mediated transformation is one of the most commonly used genetic transformation method that involves transfer of foreign genes into target plants. Agroinfiltration, an Agrobacterium-based transient approach and the breakthrough discovery of CRISPR/Cas9 holds trending stature to perform targeted and efficient genome editing (GE). The predominant feature of agroinfiltration is the abolishment of Transfer-DNA (T-DNA) integration event to ensure fewer biosafety and regulatory issues besides showcasing the capability to perform transcription and translation efficiently, hence providing a large picture through pilot-scale experiment via transient approach. The direct delivery of recombinant agrobacteria through this approach carrying CRISPR/Cas cassette to knockout the expression of the target gene in the intercellular tissue spaces by physical or vacuum infiltration can simplify the targeted site modification. This review aims to provide information on Agrobacterium-mediated transformation and implementation of agroinfiltration with GE to widen the horizon of targeted genome editing before a stable genome editing approach. This will ease the screening of numerous functions of genes in different plant species with wider applicability in future.

Experimental Study on Utilization FGD Byproducts in Building Bricks

2010 ◽  
Vol 150-151 ◽  
pp. 753-757 ◽  
Author(s):  
Xiong Hao Li ◽  
Yong Jie Xue ◽  
Min Zhou
Keyword(s):  
Experimental Study ◽  
Compressive Strength ◽  
Optimum Design ◽  
Power Plants ◽  
Pilot Scale ◽  
Mixture Composition ◽  
Technical Requirement ◽  
Do No Harm ◽  
Maximum Compressive Strength ◽  
Scale Experiment

This paper discussed the feasibility of unburned and non-autoclaved, steam cured bricks prepared by FGD byproducts from coal-fired power plants. The results show that FGD byproduct, aggregates, cementious materials and water could be used to prepare bricks during the process of stir and compaction under natural cure and steam cured condition. S4 and Z2 are the optimum design mixture composition. The maximum compressive strength and saturation coefficient are 28.7 MPa and 96.7%. FGD byproducts do no harm to environment and a pilot-scale experiment demonstrates that bricks made with FGDA can meet the MU10 level bricks technical requirement.

scholarly journals Au@CoS-BiVO4 {010} Constructed for Visible-Light-Assisted Peroxymonosulfate Activation

Catalysts ◽  
2021 ◽  
Vol 11 (11) ◽  
pp. 1414
Author(s):  
Yekun Ji ◽  
Ye Zhou ◽  
Jinnan Wang ◽  
Aimin Li ◽  
Weilin Bian ◽  
...  
Keyword(s):  
Visible Light ◽  
Charge Separation ◽  
Reaction System ◽  
Light Response ◽  
Pilot Scale ◽  
Sulfate Radical ◽  
Radical Trapping ◽  
Visible Light Response ◽  
Photogenerated Electrons ◽  
Scale Experiment

A visible-light-Fenton-like reaction system was constructed for the selective conversion of peroxymonosulfate to sulfate radical. Au@CoS, when doped on monoclinic BiVO4 {010} facets, promoted spatial charge separation due to the different energy band between the m-BiVO4 {010} and {110} facets. The visible-light response of m-BiVO4 was enhanced, which was attributed to the SPR effect of Au. And the photogenerated electrons were transferred from the m-BiVO4 {010} facet to Au via a Schottky junction. Owing to higher work function, CoS was able to capture these photoelectrons with acceleration of the Co(Ⅱ)/Co(Ⅲ) redox, enhancing peroxymonosulfate conversion to sulfate radical (Co2+ + HSO5−→ Co3+ + •SO4− + OH−). On the other hand, holes accumulated on m-BiVO4 {110} facets also contributed to organics oxidation. Thus, more than 95% of RhB was degraded within 40 min, and, even after five cycles, over 80% of RhB could be removed. The radical trapping experiments and EPR confirmed that both the sulfate radical and photogenerated hole were the main species for organics degradation. UV-vis DRS, photoluminescence (PL) and photoelectrochemical analyses also confirmed the enhancement of the visible-light response and charge separation. In a pilot scale experiment (PMS = 3 mM, initial TOC = 151 mg/L, reaction time = 4 h), CoS-Au-BiVO4 loaded on glass fiber showed a high mineralization rate (>60%) of practical wastewater.

CRISPR/Cas9-induced β-carotene Hydroxylase Mutation in Dunaliella Salina CCAP19/18

2021 ◽  
Author(s):  
Lina Hu ◽  
Shu ying FENG ◽  
Gaofeng Liang ◽  
Jingxia Du ◽  
Aifang Li ◽  
...  
Keyword(s):  
High Performance ◽  
Dunaliella Salina ◽  
Gene Editing ◽  
Expression System ◽  
Transformation Method ◽  
Practical Significance ◽  
Exon 1 ◽  
Targeted Gene Editing ◽  
Foreign Genes ◽  
Β Carotene

Abstract Dunaliella salina (D. salina) has been exploited as a novel expression system for the field of genetic engineering. However, owing to the low or inconsistent expression of target proteins, it has been greatly restricted to practical production of recombinant proteins. Since the accurate gene editing function of CRISPR/Cas system, β-carotene hydroxylase gene was chosen as an example to explore D. salina application with the purpose of improving expression level of foreign genes. In this paper, based on pKSE401 backbone, three CRISPR/Cas9 binary vectors were constructed to targeting exon 1 and 3 of the β-carotene hydroxylase of D. salina CCAP19/18 (Dschyb). D. salina mutants were obtained by salt gradient transformation method, and the expression of Dschyb gene were identified through real-time fluorescent quantitative PCR. Moreover, carotenoids content was analyzed by high-performance liquid chromatography at different time points after high intensity treatment. Compared with wild type strains, the β-carotene levels of mutants showed a significant increase, nearly up to 1.4 μg/ml, and the levels of zeaxanthin decreased to various degrees in mutants. All the results provide a compelling evidence for targeted gene editing in D. salina. This study gave a first successful gene editing of D. salina which has a very important practical significance for increasing carotene yield and meeting realistic industry demand. Furthermore, it provides an approach to overcome the current obstacles of D. salina, and then gives a strong tool to facilitates the development and application of D. salina system.

scholarly journals Efficient isolation of protoplasts from rice calli with pause points and its application in transient gene expression and genome editing assays

Plant Methods ◽  
2020 ◽  
Vol 16 (1) ◽  
Author(s):  
Snigdha Poddar ◽  
Jaclyn Tanaka ◽  
Jamie H. D. Cate ◽  
Brian Staskawicz ◽  
Myeong-Je Cho
Keyword(s):  
Gene Expression ◽  
Suspension Culture ◽  
Genome Editing ◽  
Time Course ◽  
Transient Gene Expression ◽  
Protoplast Isolation ◽  
Transient Transfection ◽  
Study Gene Expression ◽  
Rice Calli

Abstract Background An efficient in vivo transient transfection system using protoplasts is an important tool to study gene expression, metabolic pathways, and multiple mutagenesis parameters in plants. Although rice protoplasts can be isolated from germinated seedlings or cell suspension culture, preparation of those donor tissues can be inefficient, time-consuming, and laborious. Additionally, the lengthy process of protoplast isolation and transfection needs to be completed in a single day. Results Here we report a protocol for the isolation of protoplasts directly from rice calli, without using seedlings or suspension culture. The method is developed to employ discretionary pause points during protoplast isolation and before transfection. Protoplasts maintained within a sucrose cushion partway through isolation, for completion on a subsequent day, per the first pause point, are referred to as S protoplasts. Fully isolated protoplasts maintained in MMG solution for transfection on a subsequent day, per the second pause point, are referred to as M protoplasts. Both S and M protoplasts, 1 day after initiation of protoplast isolation, had minimal loss of viability and transfection efficiency compared to protoplasts 0 days after isolation. S protoplast viability decreases at a lower rate over time than that of M protoplasts and can be used with added flexibility for transient transfection assays and time-course experiments. The protoplasts produced by this method are competent for transfection of both plasmids and ribonucleoproteins (RNPs). Cas9 RNPs were used to demonstrate the utility of these protoplasts to assay genome editing in vivo. Conclusion The current study describes a highly effective and accessible method to isolate protoplasts from callus tissue induced from rice seeds. This method utilizes donor materials that are resource-efficient and easy to propagate, permits convenience via pause points, and allows for flexible transfection days after protoplast isolation. It provides an advantageous and useful platform for a variety of in vivo transient transfection studies in rice.

Effect of Superphosphate as Additive on Nitrogen and Carbon Losses during Pig Manure Composting

2013 ◽  
Vol 295-298 ◽  
pp. 1675-1679 ◽  
Author(s):  
Yi Ming Luo ◽  
De Gang Xu ◽  
Guo Xue Li
Keyword(s):  
Total Nitrogen ◽  
Ammonia Volatilization ◽  
Pilot Scale ◽  
N2o Emission ◽  
Molar Ratio ◽  
Pig Manure ◽  
Initial Carbon ◽  
P Content ◽  
Scale Experiment ◽  
Carbon Losses

A pilot scale experiment of composting in rotting boxes used pig manure with cornstalks was carried out to study the effects of superphosphate on nitrogen and carbon losses including gas emissions. Besides control, there were five amended treatments with superphosphate addition (counted by P content) at 0.05, 0.10, 0.15, 0.20 and 0.25 molar ratio of initial nitrogen. The results indicated that superphosphate addition decreased ammonia volatilization, total nitrogen and carbon losses in composting. Total nitrogen losses of superphosphate-amended treatments were reduced by 9.3%, 14.6%, 30.1%, 45.8% and 71.5%, respectively. About 0.8% to 1.2% of initial nitrogen lost in the form of N2O emission, and the CH4 emission accounted for less than 0.3% of initial carbon. More than 0.15 molar ratio of superphosphate in compost materials caused considerable adverse effects on degradation of organic matter.

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