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Analysis of KED-mediated wound response to biotic stress and mechanical damage in tomato plants (Solanum lycopersicum)
Title
Analysis of KED-mediated wound response to biotic stress and mechanical damage in tomato plants (Solanum lycopersicum).
Creator
Nifakos, Nicholas, Zhang, Xing-Hai, Florida Atlantic University, Department of Biological Sciences, Charles E. Schmidt College of Science
Abstract/Description
Lysine-rich KED was previously identified from wounded tobacco (Nicotiana tabacum) leaves before the alignment of protein sequences between NtKED (Nicotiana tabacum KED) and SlKED (Solanum lycopersicum KED) were discovered to display 55.1% identity. Using previously generated SlKED knockout plants by CRISPR/Cas9, we performed biological assays, to investigate the role of KED in wound response to biotic and abiotic stress. Previous studies implied that the KED gene functions as a role in the...
Show moreLysine-rich KED was previously identified from wounded tobacco (Nicotiana tabacum) leaves before the alignment of protein sequences between NtKED (Nicotiana tabacum KED) and SlKED (Solanum lycopersicum KED) were discovered to display 55.1% identity. Using previously generated SlKED knockout plants by CRISPR/Cas9, we performed biological assays, to investigate the role of KED in wound response to biotic and abiotic stress. Previous studies implied that the KED gene functions as a role in the wound-induced mechanism, as well as suggested that it may also function in the plant defense system against biotic stress and insect herbivory. The results from bioassays using tobacco hornworm (Manduca sexta) have proven inconclusive thus far. Expression of KED is induced not only by mechanical wounding but also by touching such as brushing the leaves, indicating that this gene is sensitive to subtle environmental signal and may be involved in defense response against abiotic stress. To further investigate the KED gene’s role in the plant defense system, biological assays using both specialist and generalist herbivores, transcription analysis using various phytohormone mutant plants, and Evans blue cellular damage assays were performed. Our findings imply that the KED gene does not seem to have a long-term effect on insect herbivory but may have a shortterm anti-feeding effect against insect herbivores. Results from the Evans blue membrane damage assay indicate the KED gene may provide some benefit to mechanically damaged plants in a short-term period post-wounding of leaf tissues. Using the SlKED knockout as genetic tool, we conclude that this gene does not confer resistance to insect herbivores over a long-term but seems to provide a beneficial defense response in the short-term. Our membrane damage assay results also imply that this gene may be involved in membrane stabilization and repair of cellular damage after mechanical wounding.
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Date Issued
2022
PURL
http://purl.flvc.org/fau/fd/FA00014001
Subject Headings
Plant gene expression, Solanum lycopersicum, Tomatoes--Effect of stress on
Format
Document (PDF)
Analysis of SLKED gene expression in CRISPR/Cas9-mediated gene knockouts in Tomato (Micro-Tom)
Title
Analysis of SLKED gene expression in CRISPR/Cas9-mediated gene knockouts in Tomato (Micro-Tom).
Creator
Vichyavichien, Paveena, Zhang, Xing-Hai, Florida Atlantic University, Charles E. Schmidt College of Science, Department of Biological Sciences
Abstract/Description
Clustered regularly interspaced short palindromic repeats (CRISPR/CRISPR-associated (Cas) protein system, CRISPR/Cas9, uses single-guide RNA to guide Cas9 to the target site for genome editing. In this study, the CRISPR/Cas9 system was used to knockout KED in tomato (Solanum lycopersicum). KED was first identified while screening the wounded tobacco (Nicotiana tabacum) leaves. We found that alignment of the protein sequence of SlKED (Solanum lycopersicum KED) and NtKED (Nicotiana tabacum KED)...
Show moreClustered regularly interspaced short palindromic repeats (CRISPR/CRISPR-associated (Cas) protein system, CRISPR/Cas9, uses single-guide RNA to guide Cas9 to the target site for genome editing. In this study, the CRISPR/Cas9 system was used to knockout KED in tomato (Solanum lycopersicum). KED was first identified while screening the wounded tobacco (Nicotiana tabacum) leaves. We found that alignment of the protein sequence of SlKED (Solanum lycopersicum KED) and NtKED (Nicotiana tabacum KED) showed 55.1% identity. To investigate, we generated SlKED knockout tomato plants with a single base pair deletion, a five base pair deletion and a three base pair deletion with a single base pair insertion. We performed wounding assays and analyzed gene expression and found that the wounded SlKED knockout plant showed no gene induction. Furthermore, the biological assay results revealed that the tobacco hornworm (Manduca sexta) gained more mass when fed on the SlKED knockout plant. Our studies show that the KED gene plays a role in wound-induced mechanism and suggested it may involve in the plant defense system against biological stress and insect feeding.
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Date Issued
2019
PURL
http://purl.flvc.org/fau/fd/FA00013275
Subject Headings
Genome editing, Gene expression, CRISPR/Cas9, CRISPR-associated protein 9
Format
Document (PDF)
Analysis of Tardigrade Damage Suppressor Protein (Dsup) Expressed in Tobacco
Title
Analysis of Tardigrade Damage Suppressor Protein (Dsup) Expressed in Tobacco.
Creator
Kirke, Justin, Zhang, Xing-Hai, Florida Atlantic University, Department of Biological Sciences, Charles E. Schmidt College of Science
Abstract/Description
DNA damage is one of the most harmful stress inducers in living organisms. Studies have shown that exposure to high doses of various types of radiation cause DNA sequence changes (mutation) and disturb protein synthesis, hormone balance, leaf gas exchange and enzyme activity. Recent discovery of a protein called Damage Suppressor Protein (Dsup), found in the tardigrade species Ramazzotius varieornatus, has shown to reduce the effects of radiation damage in human cell lines. We have generated...
Show moreDNA damage is one of the most harmful stress inducers in living organisms. Studies have shown that exposure to high doses of various types of radiation cause DNA sequence changes (mutation) and disturb protein synthesis, hormone balance, leaf gas exchange and enzyme activity. Recent discovery of a protein called Damage Suppressor Protein (Dsup), found in the tardigrade species Ramazzotius varieornatus, has shown to reduce the effects of radiation damage in human cell lines. We have generated multiple lines of tobacco plants expressing the Dsup gene and preformed numerous tests to show viability and response of these transgenic plants when exposed to mutagenic chemicals, UV radiation and ionizing radiation. We have also investigated Dsup function in association to DNA damage and repair in plants by analyzing the expression of related genes using RT-qPCR. We have also analyzed DNA damage from X-ray and UV treatments using an Alkaline Comet Assay. This project has the potential to help generate plants that are tolerant to more extreme stress environments, particularly DNA damage and mutation, unshielded by our atmosphere. The possibility of growing plants accompanying human space travel and extraterrestrial colonization inspires our imagination. Extremotolerant tardigrade genes such as Dsup may be a valuable avenue in helping to cultivate crops in these future endeavors.
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Date Issued
2019
PURL
http://purl.flvc.org/fau/fd/FA00013384
Subject Headings
DNA damage, DNA Damage--radiation effects, Tardigrada, DNA Repair, Transgenic plants
Format
Document (PDF)
Expression of a Dehydrin from the Polar Plant Cerastium arcticum in Transgenic Tobacco
Title
Expression of a Dehydrin from the Polar Plant Cerastium arcticum in Transgenic Tobacco.
Creator
Hill, William, Zhang, Xing-Hai, Florida Atlantic University, Charles E. Schmidt College of Science, Department of Biological Sciences
Abstract/Description
Water scarcity induced by drought, temperature, and salinity has plagued agricultural sustainability in recent years with unprecedented revenue losses, raising concerns for worldwide food security. Recent studies have revealed unique botanical response mechanisms to combat water related stress, namely the expression of proteins known as the dehydrins. Dehydrin proteins have been shown to serve various intracellular protective functions. The gene for a SK5 type dehydrin from the arctic plant...
Show moreWater scarcity induced by drought, temperature, and salinity has plagued agricultural sustainability in recent years with unprecedented revenue losses, raising concerns for worldwide food security. Recent studies have revealed unique botanical response mechanisms to combat water related stress, namely the expression of proteins known as the dehydrins. Dehydrin proteins have been shown to serve various intracellular protective functions. The gene for a SK5 type dehydrin from the arctic plant Cerastium arcticum (CaDHN) was introduced into tobacco plants and water deficit tolerance was evaluated. Plants overexpressing CaDHN displayed improved tolerance to salt stress, but no improvement was observed under drought stress.
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Date Issued
2015
PURL
http://purl.flvc.org/fau/fd/FA00004505, http://purl.flvc.org/fau/fd/FA00004505
Subject Headings
Cerastium., Plant bioitechnology., Plants, Effect of stress on., Crops, Effect of stress on., Plant gene expression., Plant proteins., Recombinant proteins.
Format
Document (PDF)
Nitrate Use Efficiency In Tobacco Plants Constitutively Expressing A Maize Nitrate Transporter ZmNRT2.1
Title
Nitrate Use Efficiency In Tobacco Plants Constitutively Expressing A Maize Nitrate Transporter ZmNRT2.1.
Creator
Cruz, Jessica, Zhang, Xing-Hai, Florida Atlantic University, Charles E. Schmidt College of Science, Department of Biological Sciences
Abstract/Description
The NRT2 (high affinity nitrate transporter 2) family is a part of the iHATS (inducible high affinity system) that studies have shown is responsible for the influx of nitrate into the plant cell after provision of nitrate. The ZmNRT2.1 from Zea mays was constitutively expressed in Nicotiana tabacum. To assess how over-expression of this foreign NRT2.1 affects nitrate influx by plants, nitrate content in leaf and root tissue, gene expression, and vegetal growth were analyzed in media with...
Show moreThe NRT2 (high affinity nitrate transporter 2) family is a part of the iHATS (inducible high affinity system) that studies have shown is responsible for the influx of nitrate into the plant cell after provision of nitrate. The ZmNRT2.1 from Zea mays was constitutively expressed in Nicotiana tabacum. To assess how over-expression of this foreign NRT2.1 affects nitrate influx by plants, nitrate content in leaf and root tissue, gene expression, and vegetal growth were analyzed in media with deficient or high nitrate concentrations (0.1, 1, or 10 mM). Compared to wild type plants: the transgenic lines had a significantly larger fresh weight in all nitrate conditions; primary root length was significantly longer in the 0.1 and 1 mM nitrate conditions; both the fresh weight and the primary root length were significantly higher when 50 mM NaCl was applied as a stress factor to medias containing 0.1 and 10 mM nitrate.
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Date Issued
2015
PURL
http://purl.flvc.org/fau/fd/FA00004492, http://purl.flvc.org/fau/fd/FA00004492
Subject Headings
Nitrogen--Fixation., Nitrogen-fixing plants--Metabolism., Crops and nitrogen., Field crops--Genetic engineering., Plants--Effect of nitrogen on., Soil microbiology.
Format
Document (PDF)
Targeted Gene Knock-out Via Promoter Tagging in Tomato (Lycopersicon Esculentum) and Tobacco (Nicotiana Tabacum).
Title
Targeted Gene Knock-out Via Promoter Tagging in Tomato (Lycopersicon Esculentum) and Tobacco (Nicotiana Tabacum).
Creator
Kazy, Alia, Zhang, Xing-Hai, Florida Atlantic University, Charles E. Schmidt College of Science, Department of Biological Sciences
Abstract/Description
Many different ways to create mutants have been established. This research demonstrates yet another variation of the promoter tagging technique that allows for a single step selection of the putative transgenic plants that have a mutation in constitutively expressed genes. While tomato transformants have not yet been convincingly confirmed, tobacco transformation resulted in seven transgenic lines showing resistance to high concentrations of kanamycin. Two transgenic lines were further...
Show moreMany different ways to create mutants have been established. This research demonstrates yet another variation of the promoter tagging technique that allows for a single step selection of the putative transgenic plants that have a mutation in constitutively expressed genes. While tomato transformants have not yet been convincingly confirmed, tobacco transformation resulted in seven transgenic lines showing resistance to high concentrations of kanamycin. Two transgenic lines were further investigated and three putative promoters isolated. Transient expression analysis of leaves transformed by particle bombardment with vectors carrying beta-glucuronidase gene driven by these putative promoters suggests two of them to be functional. Further investigation is needed to confirm the expression in the stably transformed plants as well as cloning of the genes downstream of the functional promoters and research of their functions.
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Date Issued
2007
PURL
http://purl.flvc.org/fau/fd/FA00000779
Subject Headings
Gene mapping--Methodology, Gene mapping--Data processing, Gene expression, Microbial genetics, Computational biology
Format
Document (PDF)
The expression and analysis of a lysine-rich wound-response protein in tomato plants.
Title
The expression and analysis of a lysine-rich wound-response protein in tomato plants.
Creator
Kaplan, Noah, Zhang, Xing-Hai, Florida Atlantic University, Charles E. Schmidt College of Science, Department of Biological Sciences
Abstract/Description
Understanding the genetic regulation of the response to wounding and wound healing in fruiting plants is imperative to maintaining agricultural sustainability, preserving the quality of food supplies, and ensuring the economic viability of agriculture. Many genes are known to be induced by wounding, providing both structural repair and defense. The KED gene in tobacco (Nicotiana tabacum) has been shown to be induced by wounding. We have identified its homologue gene in tomato (Solanum...
Show moreUnderstanding the genetic regulation of the response to wounding and wound healing in fruiting plants is imperative to maintaining agricultural sustainability, preserving the quality of food supplies, and ensuring the economic viability of agriculture. Many genes are known to be induced by wounding, providing both structural repair and defense. The KED gene in tobacco (Nicotiana tabacum) has been shown to be induced by wounding. We have identified its homologue gene in tomato (Solanum lycopersicum) that we named SlKED. We have analyzed gene expression pattern of SlKED through tomato growth and development and in response to wounding as well as hormonal and inhibitor treatments. We found that the plant hormone ethylene played a major role in the expression of SlKED. To further identify evidence for physiological and transductional functions of KED and SlKED, the tobacco KED gene was introduced to tomato and overexpressed by the fruit tissue-active PUN1 promoter from pepper (Capsicum annuum,). The expression of this gene was compared to the expression of the native SlKED gene and other known wound response genes in both the wild-type and transgenic tomato plants. The upregulation of the native SlKED gene by wounding was significantly muted in the tobacco KED-expressing transgenic plants. The expression of other genes known to be associated with wound response transduction pathways was also altered. Our studies implicate the KED gene in defense mechanisms for mechanical stress in tomato plants.
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Date Issued
2016
PURL
http://purl.flvc.org/fau/fd/FA00004773, http://purl.flvc.org/fau/fd/FA00004773
Subject Headings
Wound healing., Wounds and injuries--Genetic aspects., Plant gene expression., Plant genetic regulation., Nanostructured materials--Physiological effect., Biomedical engineering.
Format
Document (PDF)