Document Type

Thesis - Open Access

Award Date


Degree Name

Master of Science (MS)


Biology and Microbiology

First Advisor

Senthil Subramanian


ARFs, auxin, AuxRE, cloning, infiltration, nodule development


Nitrogen (N) is an important plant nutrient but its abundance in the soil is not sufficient for profitable crop production. N input in the form of chemical fertilizers helps fill this need; however, an alternative for fertilizers is an immediate need due to the environmental pollution resulting from excessive use of fertilizers. Leguminous plants such as soybean (Glycine max) that form root nodules through symbiotic association with N-fixing rhizobia and therefore need little or no chemical N fertilizers. The plant hormone auxin plays a crucial role in determining the number of nodules and their rate of maturity in soybean. Auxin action is mediated by a group of transcriptional factors named auxin response factors (ARFs) that bind conserved DNA elements named auxin response elements (AuxREs) and regulate gene expression. One of the soybean ARFs, GmARF16-2, is believed to regulate auxinresponsive gene expression during soybean nodule development; however, its mechanism of action is still unclear. This project seeks to resolve the mechanism of GmARF16-2 action using transient expression with agroinfiltration method in Nicotiana benthamiana. Previous studies have characterized GmARF16-2 as a repressor while GmARF8a as an activator ARF. We hypothesized that the balance between GmARF16-2 and GmARF8a plays a role in determining the nature and extent of auxin-responsive gene expression. In this study, GmARF16-2 transactivated direct repeat AuxRE (DR5:GUS) suggesting GmARF16-2 can potentially act as an activator ARF; however, it failed to transactivate evert repeat AuxREs (ER7GG and ER7TC). In contrast, GmARF8a, was able to transactivate evert repeat AuxREs but not with DR5:GUS. It appears that monomeric ARFs not capable of dimerization are more likely to bind and transactivate direct repeat or single-copy AuxREs and ARFs capable of dimerization are more likely to bind and transactivate evert repeat AuxREs. Therefore, the repertoire of genes regulated by each ARF is likely to be determined by the orientation and sequence of the AuxREs present in target genes. A better understanding of how auxin regulates gene expression during nodule development can help devise strategies to optimize the number of nodules and their rate of maturity to enhance nitrogen fixation.

Library of Congress Subject Headings

Soybean -- Genetics.
Plant gene expression.
Genetic regulation.
Plant hormones.



Number of Pages



South Dakota State University


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Available for download on Thursday, January 20, 2022