Molecular farming
Seyed Javad Davarpanah; Majid Dana; Gholamreza Bakhshi Khaniki; Amir Abbas Mokhtarieh
Abstract
Laccases are a group of glycoproteins which can oxidize a wide range of compounds with various biological activities and industrial applications in food and beverage, pharmaceutical, textile, and even military-related industries. Regarding this enzyme structure and the ability of plant protein production ...
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Laccases are a group of glycoproteins which can oxidize a wide range of compounds with various biological activities and industrial applications in food and beverage, pharmaceutical, textile, and even military-related industries. Regarding this enzyme structure and the ability of plant protein production machinery for protein glycosylation, a construct consisting of fungal laccaseII under control of root-specific mannopine synthase promoter and tobacco etch virus translation enhancer was designed for tobacco transformation to be used in phytoremediation. N-terminal addition of acidic tobacco endochitinase Q to Laccase directs its apoplastic secretion. Putative laccase agrobacterium-mediated transformants were confirmed using polymerase chain reaction. Semi-quantitative PCR of roots and leaves of putative transformants showed differential expression of the laccase gene at the transcriptomic level resulting from the differential function of bacterial mannopine synthase promoter. Western blotting results confirmed production of mature protein in roots which also confirms the correct function of signal peptide and secretion of this enzyme into the apoplastic space of roots. Regarding their application for protein production or phytoremediation transgenics of interest should be screened based on protein concentration and enzyme activity.
Genetic Engineering and Gene Transformation
Katayoun Zamani
Volume 8, Issue 22 , September 2018, , Pages 65-79
Abstract
Transient expression is widely used as an alternative method for stable transformation of plants, especially cereals, trees and recalcitrants. In this method, multiple copies of transgene are introduced in to the plant cell and then are transcribed and translated. As introduced gene is not integrated ...
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Transient expression is widely used as an alternative method for stable transformation of plants, especially cereals, trees and recalcitrants. In this method, multiple copies of transgene are introduced in to the plant cell and then are transcribed and translated. As introduced gene is not integrated in the genome, its expression is not affected by epigenetic factors and the location of insertion. Transient expression is a convenient method for functional analysis of genes including overexpression, silencing, gene expression networks, protein localization, promoter analysis, identification of biosynthesis pathways and so on. Additionally, this method is applied in biotechnological industries like production of recombinant proteins. Transient transformation is performed by different methods, however, many of them have overlap. So, a combination of these methods can be used for transient expression. In this review, various transient transformation techniques, their advantages and disadvantages, latest progress regarding transient expression in model and crop plants and potential and limitations regarding the application of transient expression technique in science and industry will be discussed.
Genetic Engineering and Gene Transformation
Motahareh Mohsen Por; Masoud Tohid Far
Volume 1, Issue 1 , March 2012, , Pages 35-48
Abstract
A system was designed using E. coli heat shock promoter (groE) in plastid vector and a hybrid plant/bacteria sigma factor was constructed under control of a tissue specific promoter. This system was designed for overcome to deleterious effects on plant growth and fertility that may be caused by transgene ...
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A system was designed using E. coli heat shock promoter (groE) in plastid vector and a hybrid plant/bacteria sigma factor was constructed under control of a tissue specific promoter. This system was designed for overcome to deleterious effects on plant growth and fertility that may be caused by transgene overexpression. So that hybrid sigma factors contained N- terminal motives of tobacco sigma factors including chloroplast signal peptide and RNA polymerase interaction domains, composed by C-terminal motif of E. coli sigma32 that able to recognition and binding to groE promoter. Then this gene, HSig, was cloned in Agrobacterium vector after adding regulatory elements. The result vector was used for transformation of an Iranian variety of tobacco. Detection of transgenic plants was performed by PCR, southern blot and RT-PCR analysis. The Hsig gene expression and its targeting to plastid was confirmed after transformation of tobacco chloroplast using gene gun technique for targeting of green florescent protein (GFP) under control of groE promoter using pFNGi vector into transgenic HSig explants. We hope that the system that was designed and constructed in this study for GFP expression in chloroplast genome, be able to apply in molecular farming for expression of any other desired genes instead of GFP for specific gene expression in chloroplast.
