scholarly journals Expression of erythropoietin in Indian tetraploid potato variety

F1000Research ◽  
2012 ◽  
Vol 1 ◽  
pp. 26
Author(s):  
Priti N Desai ◽  
Harish Padh
Keyword(s):  
Transgenic Plants ◽  
Mammalian Cells ◽  
Capital Investment ◽  
Transgene Expression ◽  
Present Report ◽  
Expression System ◽  
Expression Vectors ◽  
Pcr Analysis ◽  
Tissue Specific ◽  
Tissue Specific Promoter

With the advent of protein-based biotech drugs in the market, the quest for the “perfect” protein expression system, which is both economical and effective, has come into focus. Currently bacteria, yeast, insect cells, mammalian cells, transgenic animal and transgenic plants are widely used for the expression of therapeutic proteins. Among these, transgenic plants provide advantages in terms of low production cost, lower capital investment in infrastructure, and suitable post-translational modifications. The major limitation of plants as an expression host is the low level of transgene expression. To increase the expression of heterologous proteins in plants, a number of approaches have been used. One of the approaches is to increase the transgene expression by using tissue-specific promoter(s) which can concentrate the protein of interest in targeted tissues and, thus, prove advantageous in downstream purification. In the present report, a protocol for expression of heterologous protein erythropoietin in potato tuber using patatin, the tuber-tissue-specific promoter, was standardized. Expression vectors for production of the erythropoietin gene under tissue-specific promoter were successfully constructed. For production of a transgenic plant, tissue culture techniques for regeneration of the whole plant from single explants were standardized. Polymerase chain reaction (PCR) analysis was performed to confirm the stable integration of the erythropoietin gene in the potato plant by using sequence-specific primers.

scholarly journals A mini review: RuBisCo small subunit as a strong, green tissue-specific promoter

2011 ◽  
Vol 63 (2) ◽  
pp. 299-307 ◽  
Author(s):  
Allah Bakhsh ◽  
Rao Qayyum ◽  
Zeeshan Shamim ◽  
Tayyab Husnain
Keyword(s):  
Gene Expression ◽  
Transgenic Plants ◽  
Genetic Engineering ◽  
Transgene Expression ◽  
Small Subunit ◽  
Tissue Specific ◽  
Rubisco Small Subunit ◽  
Green Tissue ◽  
Study Gene Expression ◽  
Tissue Specific Promoter

Plant genetic transformation is a powerful application used to study gene expression in plants. Transcriptomics has the potential to rapidly increase our knowledge of spatial and temporal gene expression and lead to new promoters for research and development. The availability of a broad spectrum of promoters with the ability to regulate the temporal and spatial expression patterns of transgenes can increase the successful application of transgenic technology. A variety of promoters is necessary at all levels of genetic engineering in plants, from basic research, to the development of economically viable crops and plant commodities, it can address legitimate concerns raised about the safety and containment of transgenic plants in the environment. Compared with temporal- or spatial-specific expression of a toxin, constitutive expression of foreign proteins in transgenic plants can cause adverse effects. The constitutive overexpression of transgenes that interferes with normal processes in a plant underscores the need for refinement of transgene expression. The development of tissue-specific promoters to drive transgene expression has helped fulfill that need. Therefore, in certain circumstances it is desirable to use expression-specific promoters which only express the foreign gene in specific plant tissues or organs. This review highlights the uses and benefits reaped by the use of green tissue-specific promoter for the RuBisCo small subunit in different crops and systems and thus establishing a broad range of tissue-specific promoters. Such plant promoters that are activated precisely when and where they are needed would be ideal for genetic engineering strategies.

scholarly journals Construction of CRISPR/Cas9 expression vectors habouring gRNA targeted on SLIAA9 gene of tomato

2020 ◽  
Vol 18 (1) ◽  
pp. 147-156
Author(s):  
Bui Manh Minh ◽  
Ha Hong Hanh ◽  
Le Thi Thu Hien ◽  
Huynh Thi Thu Hue
Keyword(s):  
Transgenic Plants ◽  
Genome Editing ◽  
Tomato Fruit ◽  
Expression System ◽  
Secondary Compounds ◽  
Expression Vectors ◽  
Tomato Plants ◽  
Normal Morphology ◽  
Transgenic Tomato Plants ◽  
Pcr Test

Tomato (Solanum lycopersicum) is a nutritious fruit containing many secondary compounds with health benefits. The formation of tomato fruit through fertilization is controlled by auxin through Aux/IAA9 and ARF8 proteins. The mutated SlIAA9 gene leads to the parthenocarpic development of fruit or seedless tomato fruit. Nowadays, the CRISPR/Cas9 genome editing system is becoming increasingly popular in modifying desired genes on plant objects. In this study, gRNAs which target on tomato SlIAA9 gene were designed and inserted into CRISPR/Cas9 vectors. In addition, two strains of A. tumefaciens harboring pRGEB31-IAA9G2 and pRGEB32-IAA9G2 vectors carrying CRISPR/Cas9 expression system towards SlIAA9 gene in tomato were successfully created. The strain of A. tumefaciens harboring pRGEB31- IAA9G2 plasmid was used to develop transgenic tomato plants from Micro-Tom variety. PCR test showed that 5/14 plants had the presence of Cas9 gene in T0 plants. The transgenic plants have a normal morphology in comparation with the controls. The evaluation of mutant efficiency, type, and stability of mutations on the SlIAA9 will be conducted on next-generation plants when the mutations are stable and segregated into descendents.

scholarly journals Rosa26 Locus Supports Tissue-Specific Promoter Driving Transgene Expression Specifically in Pig

PLoS ONE ◽  
2014 ◽  
Vol 9 (9) ◽  
pp. e107945 ◽  
Author(s):  
Qingran Kong ◽  
Tang Hai ◽  
Jing Ma ◽  
Tianqing Huang ◽  
Dandan Jiang ◽  
...  
Keyword(s):  
Transgene Expression ◽  
Tissue Specific ◽  
Rosa26 Locus ◽  
Tissue Specific Promoter

scholarly journals Endeavours of RuBisCO small subunit promoter as a tool of green tissue specific expression

2012 ◽  
Vol 48 (No. 1) ◽  
pp. 1-9 ◽  
Author(s):  
A. Bakhsh ◽  
T. Husnain
Keyword(s):  
Transgenic Plants ◽  
Genetic Engineering ◽  
Transgene Expression ◽  
Expression Patterns ◽  
Small Subunit ◽  
Foreign Gene ◽  
Specific Expression ◽  
Tissue Specific ◽  
Rubisco Small Subunit ◽  
Green Tissue

Transcriptomics has the potential to rapidly increase our knowledge of spatial and temporal gene expression and contributes to the characterization of new promoters for research and development. The successful application of transgenic technology has been further strengthened by the availability of a broad spectrum of promoters having the ability to regulate the temporal and spatial expression patterns of the transgene. A variety of promoters is necessary at all levels of genetic engineering in plants, from basic research discoveries, to development of economically viable crops and plant commodities, to addressing legitimate concerns raised about the safety and containment of transgenic plants in the environment. Compared with the temporal- or spatial-specific expression of the toxin, constitutive expression of foreign proteins in transgenic plants may cause adverse effects. Constitutive overexpression of transgenes that interfere with normal processes in a plant underscores the need for refinement of transgene expression. The development of tissue-specific promoters to drive transgene expression has helped us to fulfil that need. Therefore, in certain circumstances, it is desirable to use expression-specific promoters which express only the foreign gene in specific plant tissues or organs. This review highlights the uses and benefits reaped by researchers by using a green tissue specific promoter, RuBisCO small subunit promoter, in different crops and systems and thus establishing a broad range of tissue specific promoters. Such plant promoters that are activated precisely when and where they are needed would be ideal for genetic engineering strategies.

Delivery of HIV-1 Polyepitope Constructs Using Cationic and Amphipathic Cell Penetrating Peptides into Mammalian Cells

2020 ◽  
Vol 17 (6) ◽  
pp. 408-428 ◽  
Author(s):  
Fatemeh Namazi ◽  
Azam Bolhassani ◽  
Seyed Mehdi Sadat ◽  
Shiva Irani
Keyword(s):  
Mammalian Cells ◽  
Expression System ◽  
293T Cell ◽  
Cell Penetrating Peptides ◽  
Expression Vectors ◽  
Effective Vaccine ◽  
In Silico Studies ◽  
Cell Penetrating ◽  
Hiv 1

Background: An effective vaccine against human immunodeficiency virus 1 (HIV-1) is an important global health priority. Despite many efforts in the development of the HIV-1 vaccine, no effective vaccine has been approved yet. Recently, polyepitope vaccines including several immunogenic and conserved epitopes of HIV-1 proteins have received special attention. Methods: In this study, HIV-1 Nef, Tat, Gp160 and P24 proteins were considered for selection of immunodominant and conserved epitopes due to their critical roles in the viral life cycle and pathogenesis. At first, the Nef60-84-Nef126-144-Tat29-49-Gp16030-53-Gp160308-323-P248-151 DNA construct was designed using in silico studies. Then, the DNA construct was subcloned in pEGFP-N1 and pET- 24a (+) expression vectors and the rNef-Tat-Gp160-P24 polyepitope peptide was generated in E.coli expression system for in vitro delivery using novel cell-penetrating peptides (CPPs), LDP-NLS and CyLoP-1, in a non-covalent manner. Also, the HR9 and MPG CPPs were used to transfer the DNA construct. Results: Our results showed that the recombinant polyepitope peptide generated in Rosetta strain migrated as a clear band of ~31 kDa in SDS-PAGE. The SEM data confirmed the formation of stable nanoparticles with a size below 250 nm. MTT assay revealed that the complexes did not represent any considerable cytotoxic effect compared to untreated cells. The results of fluorescence microscopy, flow cytometry and western blotting indicated that these CPPs successfully delivered polyepitope constructs into HEK-293T cell line. Conclusion: These data suggested that these CPPs can be used as a promising approach for the development of the HIV-1 vaccine.

scholarly journals A conditional tissue-specific transgene expression system using inducible GAL4

2001 ◽  
Vol 98 (22) ◽  
pp. 12596-12601 ◽  
Author(s):  
T. Osterwalder ◽  
K. S. Yoon ◽  
B. H. White ◽  
H. Keshishian
Keyword(s):  
Transgene Expression ◽  
Expression System ◽  
Tissue Specific

scholarly journals Production of vaccines for treatment of infectious diseases by transgenic plants

2016 ◽  
Vol 107 (1) ◽  
pp. 191 ◽  
Author(s):  
Kristina LEDL ◽  
Zlata LUTHAR
Keyword(s):  
Transgenic Plants ◽  
Infectious Diseases ◽  
Disease Virus ◽  
Hepatitis B Surface Antigen ◽  
Expression System ◽  
Expression Vectors ◽  
Norwalk Virus ◽  
Mouth Disease Virus ◽  
Avian Influenza Virus H5n1 ◽  
Transformation Methods

<p>Since the first pathogen antigen was expressed in transgenic plants with the aim of producing edible vaccine in<em> </em>early 1990s, transgenic plants have become a well-established expression system for production of alternative vaccines against various human and animal infectious diseases. The main focus of plant expression systems in the last five years has been on improving expression of well-studied antigens such as porcine reproductive and respiratory syndrome (PRRSV), bovine viral diarrhea disease virus (BVDV), footh and mouth disease virus (FMDV), hepatitis B surface antigen (HBsAg), rabies G protein, rotavirus, Newcastle disease virus (NDV), Norwalk virus capsid protein (NVCP), avian influenza virus H5N1, <em>Escherichia coli</em> heat-labile enterotoxin subunit B (LT-B), cholera toxin B (CT-B), human immunodeficiency virus (HIV), artherosclerosis, ebola and anthrax. Significant increases in expression have been obtained using improved expression vectors, different plant species and transformation methods.</p>

scholarly journals Short-Term and Highly Efficient Cas9 Expression System Using Adenovirus Vector and Site-Specific Recombinase Cre

2021 ◽  
Author(s):  
Sayaka Nagamoto ◽  
Miyuki Agawa ◽  
Emi Tsuchitani ◽  
Kazunori Akimoto ◽  
Saki Kondo ◽  
...  
Keyword(s):  
Hepatitis B ◽  
Genome Editing ◽  
Expression System ◽  
Adenovirus Vector ◽  
Virus Genome ◽  
Dna Viruses ◽  
Site Specific ◽  
Tissue Specific ◽  
Tissue Specific Promoter ◽  
A Cell

Abstract Genome editing techniques such as CRISPR/Cas9 have both become common gene engineering technologies and have been applied to gene therapy. However, the problems of increasing the efficiency of genome editing and reducing off-target effects that induce double-stranded breaks at unexpected sites in the genome remain. In this study, we developed a novel Cas9 transduction system, Exci-Cas9, using an adenovirus vector (AdV). Cas9 was expressed on a circular molecule excised by the site-specific recombinase Cre and succeeded in shortening the expression period compared to AdV, which expresses the gene of interest for at least six months. As an example, we chose hepatitis B, which currently has more than 200 million carriers in the world and frequently progresses to liver cirrhosis or hepatocellular carcinoma. The efficiencies of hepatitis B virus genome disruption by Exci-Cas9 and Cas9 expression by AdV directly (Avec) were the same, about 80–90%. Furthermore, Exci-Cas9 enabled cell- or tissue-specific genome editing by expressing Cre from a cell- or tissue-specific promoter. We believe that Exci-Cas9 developed in this study is useful not only for resolving the persistent expression of Cas9, which has been a problem in genome editing, but also for eliminating long-term DNA viruses such as human papilloma virus.

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