Expression and Localization of the PabPrx86 Peroxidase from Norway Spruce (Picea abies (L.) Karst.) in Tobacco (Nicotiana benthamiana L.)

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http://urn.fi/URN:NBN:fi:hulib-202006173073
Title: Expression and Localization of the PabPrx86 Peroxidase from Norway Spruce (Picea abies (L.) Karst.) in Tobacco (Nicotiana benthamiana L.)
Author: Taniwan, Steven
Other contributor: Helsingin yliopisto, Maatalous-metsätieteellinen tiedekunta
University of Helsinki, Faculty of Agriculture and Forestry
Helsingfors universitet, Agrikultur- och forstvetenskapliga fakulteten
Publisher: Helsingin yliopisto
Date: 2020
Language: eng
URI: http://urn.fi/URN:NBN:fi:hulib-202006173073
http://hdl.handle.net/10138/316360
Thesis level: master's thesis
Degree program: Erasmus-Mundus Master Program in Plant Breeding (emPlant)
Erasmus-Mundus Master Program in Plant Breeding (emPlant)
Erasmus-Mundus Master Program in Plant Breeding (emPlant)
Specialisation: Kasvintuotantotieteet
Plant Production Sciences
Växtproduktionsvetenskaper
Abstract: Norway spruce is commonly cultivated throughout Europe, Russia, and Japan. Cultivation of Norway spruce often faces the issue of fungal diseases, one of which is cherry rust disease caused by Thekopsora areolata. The gene model MA_10g0010 encoding an uncharacterized peroxidase (PabPrx86) has previously been associated with the presence of this pathogen. The aim of this study was to describe and assay the protein produced from this gene model, observe its localization in the cell, and determine its relative expression level in different tissues of Norway spruce. Experiments were performed by isolating the full length cDNA for PabPrx86 and cloning the cDNA into destination vectors pEAQ-HT-DEST1 and pK7FWG2 leading to a hypertranslatable transcript and a C-terminal GFP fusion, respectively. The plasmid constructs were transformed to Agrobacterium tumefaciens and agro-infiltrated to Nicotiana benthamiana. In addition, the relative expression level of this gene in different spruce tissues at different times of the year was determined using the qRT-PCR method. Sequencing showed that there were two allelic variants of this gene in the spruce individual sampled for RNA. Results showed that both alleles code for a peroxidase with basic pI. Subcellular localization with the GFP tag detected that PabPrx86 protein was located out of cytoplasm, indicating that the protein was translated in the ER-ribosomes, whereas relative expression level analysis revealed that PabPrx86 was highest expressed in the bud and lateral bud in June. Peroxidases are known to relate with plant defense, but further experiments are required to determine the role of PabPrx86 in Norway spruce and what the association with T. areolata means.
Subject: Norway spruce
PabPrx86
peroxidase


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