In Situ Proteomic Analysis of Herbicide-Resistant Soybean and Hybrid Seeds via Matrix-Assisted Laser Desorption/Ionization-Mass Spectrometry Imaging

Abstract

Transgenic soybean is the commercial crop with the largest cultivation area worldwide. During transgenic soybean cultivation, exogenous genes may be transferred to wild relatives through gene flow, posing unpredictable ecological risks. Accordingly, an environmental risk assessment should focus on fitness changes and underlying mechanisms in hybrids between transgenic and wild soybeans (Glycine soja). Matrix-assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MSI) was used for in situ detection and imaging of protein changes in the seeds of transgenic herbicide-resistant soybean harboring epsps and pat genes, non-transgenic soybean, wild soybean, and their F2 hybrid. Protein data clearly distinguished wild soybeans, while the F2 seeds had protein characteristics of both parents and were distinguished from wild soybean seeds. Using UPLC-Q-TOF-MS, 22 differentially expressed proteins (DEPs) were identified, including 13 specific to wild soybean. Sucrose synthase and stress response-related DEPs were differentially expressed in parental and offspring. Differences in these may underpin the greater adaptability of the latter. MSI revealed DEP distribution in transgenic, wild, and F2 seeds. Identifying DEPs related to fitness may elucidate mechanisms underlying fitness differences among the studied varieties. Our study shows that MALDI-MSI has the potential to become a visual method for transgenic soybean analysis.