In order to investigate the molecular mechanism of Styxl1 in the regulation of sperm flagella, sperm from Styxl1+/+ and Styxl1-/- mice were collected for quantitative proteomics analysis using TMT-based liquid chromatography-tandem mass spectrometry (LC-MS/MS) approach. A total of 6429 proteins were quantified, and 1517 proteins (p < 0.05, Foldchange > 2) were differentially expressed. Among these differential proteins, 986 proteins were downregulated (65%) and 531 proteins were upregulated (35%) in Styxl1-/- sperm.