Development of a rapid detection assay for acetolactate synthase inhibitors resistance in three Amaranthus weed species through loop-mediated isothermal amplification.

BACKGROUND

The early detection of herbicide resistance in weeds is a key factor to avoid herbicide waste and improve agriculture sustainability. The aim of this work was to develop and validate an allele-specific loop-mediated isothermal amplification (AS-LAMP) assay for the quick on-site detection of the resistance-endowing point mutation Trp-574-Leu in the acetolactate synthase (ALS) gene in three widely diffused Amaranthus weed species, A. retroflexus, A. hybridus and A. tuberculatus.

RESULTS

The AS-LAMP protocol was developed on wild type and ALS-mutant plants of the three species and revealed that the amplification approach with only the primer set specific for the mutant allele (574-Leu) was the most promising. The validation and estimation of the AS-LAMP performance evaluated by comparing the results with those of the molecular marker (CAPS, cleaved amplified polymorphic sequences) indicated that while sensitivity and specificity resulted relatively high in all species (overall 100% and >65%, respectively), precision was high for Amaranthus hybridus L. and Amaranthus retroflexus L. (75% and 79%, respectively), but quite low for Amaranthus tuberculatus (Moq.) J. D. Sauer (59%). The LAMP assay was also effective on crude genomic DNA extraction, allowing the quick detection of mutant plants in field situation (on site resistance detection).

CONCLUSIONS

The proposed AS-LAMP method has proven to be a promising technique for rapid detection of resistance due to Trp-574-Leu on the two monoecious weedy Amaranthus species but resulted less effective in the genetically variable dioecious species A. tuberculatus. This article is protected by copyright. All rights reserved.

Informations de copyright

This article is protected by copyright. All rights reserved.