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.
Plant Disease and Biotechnology
Azam Badr Hadad; Farhad Nazarian Firouzabadi; Ahmad Ismaili; Hedayat Bagheri
Volume 7, Issue 19 , November 2017, , Pages 1-13
Abstract
Antimicrobial peptides are ancient and conserved molecules which are found in defense mechanisms of almost all living organisms from bacteria to animal and plant species. Identification and introduction of novel antimicrobial peptides, is a cost-effective way to improve crop plants resistance to pathogens ...
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Antimicrobial peptides are ancient and conserved molecules which are found in defense mechanisms of almost all living organisms from bacteria to animal and plant species. Identification and introduction of novel antimicrobial peptides, is a cost-effective way to improve crop plants resistance to pathogens by using recombinant DNA technology. Therefore, an expression construct containing omiganan antimicrobial encoding gene from the cytoplasmic granules of bovine neutrophils, was cloned and transferred to the tobacco leaf disk using Agrobacterium tumefaciens mediated-transformation. The presence of the antimicrobial peptide encoding gene in the genome of transgenic plants was confirmed by PCR analysis. Six putative transgenic lines and a non-transgenic control plant were selected for further molecular analysis. Total protein was extract from transgenic and non-transgenic control plants and used for antimicrobial activity assay against some human; E. coli, Salmonella, Staphylococcus, Bacillus, and plant: Xanthomonas and Pseudomonas pathogens by disc diffusion method. Results of this experiment showed that total protein extract from transgenic lines, as compared to non-transgenic plant, was significantly (P
Proteomics
Maryam Jamshidnia; Sayed Kamal Kazemitabar; Christian Lindermayr; Hamid Najafi Zarini
Volume 6, Issue 16 , March 2017, , Pages 1-12
Abstract
Recently, transient gene expression has been developed to provide a more rapid means of assessing plant tissues as a protein production platform without the labor-intensive and time-consuming process of generating stably transformed transgenic plants. This study reports the expression of HDA19 gene in ...
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Recently, transient gene expression has been developed to provide a more rapid means of assessing plant tissues as a protein production platform without the labor-intensive and time-consuming process of generating stably transformed transgenic plants. This study reports the expression of HDA19 gene in two species of tobacco plants (Nicotiana tabacum and Nicotiana bentamiana) by means of transient transformation. Specific primers were designed and used for PCR amplification and cloning of HDA19 gene in the plant expression vector pB2GW7. The recombinant construct was transferred into Agrobacterium tumefaciens strain GV3101, and was used for Agrobacterium mediated transformation of tobacco plants. The presence of the desired gene in transgenic lines was confirmed through colony PCR. The expression of the protein in transgenic lines was confirmed by immune-dot blot assay and ELISA. Although the transformation of the two species was confirmed by immune-dot blot assay and SDS-PAGE, recombinant protein production in Nicotiana tabacum plants was confirmed by ELISA and it was estimated 400 µg per gram wet weight of tobacco leaves. According to the results, this species is the appropriate host for the production of recombinant HDA19, one of the histone deacetylases, rather than Nicotiana bentamiana.
Genomics
Faramarz Hoshyardel; Reza Darvishzadeh
Volume 6, Issue 14 , August 2016, , Pages 61-72
Abstract
Tobacco (Nicotiana tabacum L.) is an industrial plant that play important role in economy of most countries. Oriental tobacco is a sun-cured, small-leafed, highly aromatic type of tobacco with limited information about its agronomic and chemical characteristics and their genetic control. This research ...
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Tobacco (Nicotiana tabacum L.) is an industrial plant that play important role in economy of most countries. Oriental tobacco is a sun-cured, small-leafed, highly aromatic type of tobacco with limited information about its agronomic and chemical characteristics and their genetic control. This research was conducted to prepare the genetic linkage map of oriental type tobacco in a recombinant inbred line population (103 lines) produced from the cross between SPT406 (male parent) and Basma seres 31 (female parent) and identification of genomic locations controlling some agronomic characteristics accompanied with chlorine content under field conditions. Markers including SSR, ISSR, REMAP and IRAP were utilized in construction of genetic linkage map. Linkage map comprising 46 markers was developed which covered 586.1 cM of tobacco genome and the average distance between two markers was 12.74 cM. Number of markers per linkage groups varied between 2 to 13. In this study, 19 QTLs with phenotypic variation between 13.2 to 40.1 were identified for characteristics including leaf length, leaf width, leaf number, plant height, internode distance, stem girth, dry leaf yield, photosynthesis value and leaf chlorine content using composite interval mapping. Co-localized QTLs were recognized in linkage groups 1, 3 and 5. Highest values of phenotypic variation achieved in the present study manifest the possibility for using identified retrotransposon and microsatellite markers in marker assisted selection programs of oriental type tobacco.
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.
