Prototype demonstration of transgenic resistance to the nematode Radopholus similis conferred on banana by a cystatin.
Cavendish banana was transformed using Agrobacterium tumefaciens to express a protein engineered rice cystatin (OcIdeltaD86) of value for control of plant parasitic nematodes. Expression for each line was under control of a constitutive promoter from the maize ubiquitin gene (UBI-1), a constitutive, chimeric promoter from the octopine and mannopine synthase genes of A. tumefaciens or a promoter from a root-preferentially expressed tubulin gene Arabidopsis (TUB-1). Lines were selected as of potential interest after 8 weeks challenge in containment if their mean R. similis/25 g roots for three sibling plants were more than 1 standard normal variate below the grand mean for all plants in c7-15 lines challenged concurrently. A total of 16 lines were selected on this basis as putative positives. Western blots confirmed that eight of these lines expressed cystatin with a mean of 0.08 +/- 0.04% tsp. All but two of 19 negatively selected lines from bioassays did not express cystatin. The mean resistance level of the confirmed positive lines was 69 +/- 17%. ELISA established the positive lines under control of UBI provided significantly higher expression levels of OcIdeltaD86 than recorded for the other two promoters. Lines of interest were confirmed as producing a transcript for OcIdeltaD86 by RT-PCR. Eight plants of one UBI promoter line expressing only 0.1 +/- 0.004% tsp as cystatin were re-challenged with R. similis and achieved a resistance of 70 +/- 10%. Subsequent repeat western blotting confirmed that this line still produced the cystatin after the trial. This is the first report of transgenic resistance against a major pest or disease of banana.
Atkinson
Howard J
HJ
Centre for Plant Sciences, University of Leeds, Leeds LS2 9JT, UK. h.j.atkinson@leeds.ac.uk
Grimwood
Sam
S
Johnston
Kate
K
Green
Jayne
J
D000818:Animals; D015891:Cystatins / Q000235:genetics* / Q000378:metabolism; D028521:Musa / Q000235:genetics* / Q000469:parasitology*; D010935:Plant Diseases / Q000469:parasitology*; D030821:Plants, Genetically Modified / Q000378:metabolism; D019076:Transgenes / Q000235:genetics; D014431:Tylenchoidea
D015891:Cystatins