Characterization of resistance to lettuce mosaic virus in Lactuca sativa

Lettuce mosaic virus (LMV) is an economically important pathogen with worldwide distribution. LMV infection in L. sativa can cause significant yield losses. Resistance to LMV in L. sativa is conferred by the recessive gene mo. We attempted to position the mo gene on the L. sativa map. The ultimate goal is a better understanding of plant-virus interactions. To do so, Random Amplified Polymorphic DNA (RAPD) markers were screened in the near isogenic lines (NILs) Vanguard and Vanguard 75. These NILs differ in the presence of the mo gene in Vanguard 75. Polymorphic markers were screened for linkage to mo in two F$sb2$ populations segregating for resistance to LMV. The F$sb2$ populations used were derived from 2 crosses, the first one between the L. sativa cultivars Dwarf 2 (resistant to LMV via the presence of mo) and Saffier and the second one between two breeding lines 87-25M-1 (momo) and 87-1090M-1 (MoMo). In order to develop a highly stringent antibody detection system to phenotype plants infected with LMV, a plasmid construct was developed which overproduces LMV coat protein. This construct will be used in the future to produce enough recombinant LMV coat protein for antibody production. To further characterize mo, a selection of cultivars resistant and susceptible to LMV according to the literature were subjected to various temperature changes to determine the environmental influences on virus movement

Study of the arbuscular mycorrhizal fungus Glomus intraradices at the molecular level

Arbuscular mycorrhizal (AM) fungi have been living in association with land plants for at least 400 million years. Because they are obligate symbionts, the study of AM symbiosis has focused primarily on its plant host and progress in the molecular biology of AM fungi has been very slow. Using two different approaches, library screening and direct PCR-based assays, genetic information of AM fungi was compared across isolates and species. This allowed the study of novel DNA regions previously unexplored in AM fungi. The following species were investigated: Glomus intraradices, Glomus mosseae, Gigaspora margarita, Scutellospora calospora, Acaulospora scrobiculata and Entrophosphora colombiana. In the first approach, using in vitro grown G. intraradices, a DNA extraction protocol was developed for the construction of a partial genomic library. This library was screened for the presence of microsatellite-containing loci. PCR primers were designed based on five identified loci. Two of these loci were monomorphic for all isolates and species. The second approach used a combination of degenerate and specific primers for fungal chitin synthase genes to explore the variability of this gene family in AM fungi. A total of 21 AM sequences were isolated and sequenced, covering class I and II chitin synthases. RT-PCR with G. intraradices revealed differential expression of chitin synthases in spores and mycelium, as compared to mycorrhized roots. In addition, using primers designed from a highly conserved sequence for plant resistance genes, classical PCR and RT-PCR allowed the detection of a genomic sequence and its cDNA counterpart encoding a putative serine/lysine rich protein in G. intraradices. We have thus investigated genetic variability in AM fungi in functional genes as well as in repetitive DNA regions. Study of gene expression was also possible using in-vitro grown G. intraradices

Heat shock 70 protein interaction with Turnip mosaic virus RNA-dependent RNA polymerase within virus-induced membrane vesicles

AbstractTandem affinity purification was used in Arabidopsis thaliana to identify cellular interactors of Turnip mosaic virus (TuMV) RNA-dependent RNA polymerase (RdRp). The heat shock cognate 70-3 (Hsc70-3) and poly(A)-binding (PABP) host proteins were recovered and shown to interact with the RdRp in vitro. As previously shown for PABP, Hsc70-3 was redistributed to nuclear and membranous fractions in infected plants and both RdRp interactors were co-immunoprecipitated from a membrane-enriched extract using RdRp-specific antibodies. Fluorescently tagged RdRp and Hsc70-3 localized to the cytoplasm and the nucleus when expressed alone or in combination in Nicotiana benthamiana. However, they were redistributed to large perinuclear ER-derived vesicles when co-expressed with the membrane binding 6K-VPg-Pro protein of TuMV. The association of Hsc70-3 with the RdRp could possibly take place in membrane-derived replication complexes. Thus, Hsc70-3 and PABP2 are potentially integral components of the replicase complex and could have important roles to play in the regulation of potyviral RdRp functions

Phytogeographic and genetic variation in Sorbus, a traditional antidiabetic medicine—adaptation in action in both a plant and a discipline

Mountain ash (Sorbus decora and S. americana) is used by the Cree Nation of the James Bay region of Quebec (Eeyou Istchee) as traditional medicine. Its potential as an antidiabetic medicine is thought to vary across its geographical range, yet little is known about the factors that affect its antioxidant capacity. Here, we examined metabolite gene expression in relation to antioxidant activity, linking phytochemistry and medicinal potential. Samples of leaf and bark from S. decora and S. americana were collected from 20 populations at four different latitudes. Two genes known to produce antidiabetic substances, flavonol synthase and squalene synthase, were analyzed using quantitative real time PCR. Gene expression was significantly higher for flavonol synthase compared to squalene synthase and increased in the most Northern latitude. Corresponding differences observed in the antioxidant capacity of ethanolic extracts from the bark of Sorbus spp. confirm that plants at higher latitudes increase production of stress-induced secondary metabolites and support Aboriginal perceptions of their higher medicinal potential. Modern genetic techniques such as quantitative real time PCR offer unprecedented resolution to substantiate and scrutinise Aboriginal medicinal plant perception. Furthermore, it offers valuable insights into how environmental stress can trigger an adaptive response resulting in the accumulation of secondary metabolites with human medicinal properties

Distributed under Creative Commons CC-BY 4.0 Phytogeographic and genetic variation in Sorbus, a traditional antidiabetic medicine-adaptation in action in both a plant and a discipline

ABSTRACT Mountain ash (Sorbus decora and S. americana) is used by the Cree Nation of the James Bay region of Quebec (Eeyou Istchee) as traditional medicine. Its potential as an antidiabetic medicine is thought to vary across its geographical range, yet little is known about the factors that affect its antioxidant capacity. Here, we examined metabolite gene expression in relation to antioxidant activity, linking phytochemistry and medicinal potential. Samples of leaf and bark from S. decora and S. americana were collected from 20 populations at four different latitudes. Two genes known to produce antidiabetic substances, flavonol synthase and squalene synthase, were analyzed using quantitative real time PCR. Gene expression was significantly higher for flavonol synthase compared to squalene synthase and increased in the most Northern latitude. Corresponding differences observed in the antioxidant capacity of ethanolic extracts from the bark of Sorbus spp. confirm that plants at higher latitudes increase production of stressinduced secondary metabolites and support Aboriginal perceptions of their higher medicinal potential. Modern genetic techniques such as quantitative real time PCR offer unprecedented resolution to substantiate and scrutinise Aboriginal medicinal plant perception. Furthermore, it offers valuable insights into how environmental stress can trigger an adaptive response resulting in the accumulation of secondary metabolites with human medicinal properties