Graduation Term
Spring 2026
Degree Name
Master of Science (MS)
Department
School of Biological Sciences
Committee Chair
John Sedbrook
Committee Member
Tom Hammond
Committee Member
Viktor Kirik
Abstract
Field pennycress (Thlaspi arvense L.; pennycress) is a winter hardy weed that grows throughout the U.S. Midwest and other temperate regions around the world. Pennycress has undergone many significant genetic changes to be domesticated as a winter-annual oilseed crop. As an intermediate crop grown from fall to spring, pennycress offers the ecological benefits of winter cover crops while also producing high-value seeds rich in oil and protein. The extracted seed oil is an attractive feedstock for renewable diesel and sustainable aviation fuel while the crushed seed after oil extraction (the seed meal) has utility as a high protein animal feed. However, wild-type pennycress seed meal suffers from high levels of anti-nutritional glucosinolates, ultimately limiting the amounts that can be used in animal feed.
To combat the high seed glucosinolate content, our lab has generated numerous mutants through CRISPR-Cas9 mutagenesis targeting sinigrin production and transportation genes. Sinigrin serves as the foremost glucosinolate produced in pennycress seeds acting as both a defensive compound and sulfur storage. The glucosinolate mutations confer a range of sinigrin content reductions but in many instances have the tradeoff of negatively impacting plant growth possibly due to the reduced bioavailability of sulfur.
To understand if the altered growth phenotypes observed with some of the reduced glucosinolate mutant lines were due to reduced sulfur availability or instead gene expression differences directly affecting plant growth and stress resilience, aop2, hag1hag3, myc3hag1, and myc3hag3 single and double mutants were grown on soils supplemented with varying concentrations of the fertilizers, ammonium sulfate and ammonium nitrate, in an attempt to rescue the mutant phenotypes. Ultimately, I found the fertilizer supplementation was ineffective at returning the mutant lines to a healthy state and often resulted in elevated concentrations of sinigrin within seed tissues, suggesting the altered growth of many if not all of the mutants was due to the mutations directly altering growth and not just sulfur availability. This is not surprising given that HAG1, HAG3, and MYC3 encode transcription factors influencing jasmonic acid and salicylic acid stress response pathways.
My thesis work also involved producing new pennycress reduced glucosinolate mutant lines by targeting CRISPR-Cas9 knockout of UMAMIT29 and UMAMIT30 which encode two glucosinolate transporters shown in Arabidopsis to export glucosinolates from source vegetative tissues to sink seed tissues. Single and double mutations were induced in the Spring 32-10 and tt8.ARV1 pennycress backgrounds. Quantification of plant growth rates and leaf glucosinolate content revealed limited differences between mutant lines and wild type. It was somewhat surprising the knockout mutations did not lead to increases in leaf glucosinolate content given that they presumably block transport out of these sink tissues. The lack of negative effects on plant growth was also somewhat surprising given that knockout of GTR glucosinolate transporters cause stunted growth. Quantification of seed glucosinolate content in umamit29 and umamit30 single and double mutants remains to be performed upon plant senescence.
Access Type
Thesis-Open Access
Recommended Citation
Bayliss, Ryan, "Integrating Fertilizer Supplementation and CRISPR-Mediated UMAMIT29/30 Mutagenesis to Optimize Low Glucosinolate Pennycress" (2026). Theses and Dissertations. 2280.
https://ir.library.illinoisstate.edu/etd/2280