Spatial monitoring of gene activity in extraradical and intraradical developmental stages of arbuscular mycorrhizal fungi by direct fluorescent in situ RT-PCR.
Gene expression profiling based on tissue extracts gives only limited information about genes associated with complex developmental processes such as those implicated in fungal interactions with plant roots during arbuscular mycorrhiza development and function. To overcome this drawback, a direct fluorescent in situ RT-PCR methodology was developed for spatial mapping of gene expression in different presymbiotic and symbiotic structures of an arbuscular mycorrhizal fungus. Transcript detection was optimized by targeting the LSU rRNA gene of Glomus intraradices and monitoring expression of a stearoyl-CoA-desaturase gene that is consistently expressed at high levels in spores, hyphae, arbuscules and vesicles. This method was further validated by localizing expression of fungal peptidylprolyl isomerase and superoxide dismutase genes, which are expressed to different extents in fungal structures. Direct fluorescent in situ RT-PCR offers new perspectives for the sensitive analysis of fungal developmental processes that occur during functional differentiation in symbiotic arbuscular mycorrhiza interactions.
Seddas
Pascale M A
PM
UMR 1088 INRA/5184 CNRS/Université de Bourgogne, Plante-Microbe-Environnement, INRA-CMSE, BP 86510, 21065 Dijon Cedex, France. pseddas@dijon.inra.fr
Arnould
Christine
C
Tollot
Marie
M
Arias
Cecilia M
CM
Gianinazzi-Pearson
Vivienne
V
D017931:DNA Primers / Q000737:chemistry / Q000235:genetics; D005456:Fluorescent Dyes / Q000737:chemistry; D005656:Fungal Proteins / Q000235:genetics; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D015966:Gene Expression Regulation, Fungal; D029902:Medicago / Q000382:microbiology; D018613:Microscopy, Confocal; D038821:Mycorrhizae / Q000235:genetics* / Q000254:growth & development; D019696:Peptidylprolyl Isomerase / Q000235:genetics; D018517:Plant Roots / Q000382:microbiology; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D013230:Stearoyl-CoA Desaturase / Q000235:genetics; D013482:Superoxide Dismutase / Q000235:genetics; D013559:Symbiosis / Q000235:genetics; D014966:Xanthenes / Q000737:chemistry
D017931:DNA Primers; D005456:Fluorescent Dyes; D005656:Fungal Proteins; D014966:Xanthenes; C034657:Texas red; D013230:Stearoyl-CoA Desaturase; D013482:Superoxide Dismutase; D019696:Peptidylprolyl Isomerase