A. palmeri is an aggressive and prolific weed species that has major ecological and economic impacts on agricultural row cropping systems. Using morphological identification to distinguish A. palmeri from other Amaranthus species is difficult, which has led to the use of genetic testing becoming the standard for Amaranthus species identification. We have developed an improved genetic test to maximize the robustness and reliability of A. palmeri identification. We assembled a large and geographically diverse panel of Amaranthus accessions and utilized Genotyping by Sequencing (GBS) to identify novel species-specific single nucleotide polymorphisms (SNPs) from these populations. These diagnostic SNPs were then used to develop a genotyping assays through Kompetative Allele Specific PCR (KASP). Performance was assessed against a validation panel of 1,250 Amaranthus individuals of 9 species.
Results/Conclusions
The end result is a set of three KASP assays for identification of A. palmeri, for use either independently or in conjunction. Assays were tested against the most robust validation panel of Amaranthus individuals in publication thus far, and demonstrated an accuracy of 99.7-99.9% on single sample validations. Diagnostic statistics and confusion matrices are presented. Additionally, these markers were tested for sensitivity by extracting A. palmeri seeds in combination with seeds from Tall Waterhemp (A. tuberculatus). All three markers are capable of reliably detecting a single A. palmeri seed in a pool of 200 Amaranthus sp. seeds (p < 0.0001). Our work represents an improvement over existing published assays in terms of sensitivity, robustness of validation, and ease of use.