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ARS Home » Midwest Area » Lexington, Kentucky » Forage-animal Production Research » Research » Publications at this Location » Publication #361211
Title: Analysis of the transcriptome following 2,4-D treatment in susceptible and tolerant red clover (Trifolium pratense) lines

Author
item ARAUJO, LUCAS - University Of Kentucky
item BARRETT, MICHAEL - University Of Kentucky
item OLSON, GENE - University Of Kentucky
item Dinkins, Randy

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 11/29/2018
Publication Date: N/A
Citation: N/A

Interpretive Summary:

Technical Abstract: Incorporation of a legume, such as red clover (Trifolium pratense), into grass-based pasture systems offers many benefits. However, available red clover lines are highly susceptible to broadleaf herbicides, including 2,4-D (2,4-dichlorophenoxyacetic acid), that are used for weed management in pastures. A novel red clover line, UK2014, was developed at the University of Kentucky through conventional breeding. UK2014 is more tolerant to 2,4-D than Kenland, a common variety used by Kentucky’s forage producers. We utilized Next Generation Sequencing (NGS) to uncover the genetic basis for the increased UK2014 2,4-D tolerance. NGS provides a novel and powerful tool to analyze the mechanisms of herbicide resistance. Specifically, this study used a NGS approach to detect differential gene expression between UK2014 and Kenland following 2,4-D treatment in a field setting. Both UK2014 and Kenland lines were grown in at the University of Kentucky’s Spindletop research farm. Plots of each line were treated with either 0 or 1.12 kg ae/ha of 2,4-D amine. Composite samples of ten leaflets from each plot were collected at 4, 24 and 72 hours after treatment (HAT). RNA sequencing was performed in the NGS Illumina HiSeq 2500 with 100 bp single reads. The reads were mapped against the T. pratense genome and read counts were obtained by the CLC workbench. The read counts were submitted to TMM normalization and analyses of variance in JMP genomics. A false discovery rate of 0.05 (FDR = 0.05) was set as the threshold for differential expression calling. The online tool AgriGO provided the gene ontology of the differentially expressed transcripts. Time of sampling (HAT) and 2,4-D treatment were the major fixed components of the variance, and cultivar displayed less of an effect. A significant number of these (997 out of 2217 contigs) was found to be upregulated in both UK2014 and Kenland following 2,4-D treatment. From these upregulated contigs, 50 were annotated as cytochromes P450 and glucosyl-transferases, enzymes commonly related to herbicide metabolism. Moreover, gene ontology of the differentially expressed contigs indicated a major relationship to biological processes such as responses to stimulus, stress, and metabolic processes. These results, as a whole, provide an indication of the processes involved in the increased UK2014 tolerance to 2,4-D, although further validation is still necessary.