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A putative Fe+3/phytosiderophore (PS) transporter isolated from oats (Avena sativa)
Author(s) -
Davis George Thomas,
Stocksdale Mark
Publication year - 2013
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.27.1_supplement.1017.2
Subject(s) - avena , genbank , complementation , citric acid , clone (java method) , transporter , mutant , saccharomyces cerevisiae , hordeum vulgare , yeast , biochemistry , chemistry , biology , microbiology and biotechnology , gene , botany , poaceae
The objectives of this project were: 1) to isolate and characterize a Fe +3 /phytosiderophore (PS) transporter from Oats ( Avena sativa ), and, 2) develop a method for the synthesis of the oat phytosiderophore avenic acid. The putative transporter's functionality will be evaluated by complementation assays in yeast ( Saccharomyces cerevisiae ) using synthetic avenic acid (Stocksdale). Total RNA was isolated from the roots of hydroponically grown, iron‐starved oats (var. Rodeo). Full‐length clones were obtained using 5′RACE and RT/PCR. Sequence analysis of the clone, AvsYS1 , (Genbank Accession# FJ477297 ) shows>;85% similarity at both the nucleotide and amino acid level to HvYS1, a functionally confirmed Fe +3 / (PS) transporter from barley ( Hordeum vulgare ). AvsYS1 was subcloned into pYES2 (Invitrogen), a galactose‐inducible shuttle vector. DDY4 ( a gift from Dr. David Eide) is a mutant S. cervisiae lacking a Fe +3 uptake ability. DDY4 will be transformed with pYES2/BS4YAT, pYES2 containing the AvsYS1 insert. To evaluate functionality, transformants will be grown in minimal media (plates and liquid) supplemented with FeCl 3 , synthetic avenic acid or complexed FeCl 3 /avenic acid. If AvsYS1 encodes a functional transporter we will be able to rescue DDY4 in a the medium is supplemented with FeCl 3 /avenic acid. This research was supported by a USDA CSREES Grant, #2007–35318