Leaf rolling is a crucial agronomic trait to consider in rice (Oryza sativaL.) breeding as it keeps the leaves upright, reducing interleaf shading and improving photosynthetic efficiency. TheSEMI-ROLLED LEAF 1(SRL1) gene plays a key role in regulating leaf rolling, as it encodes a glycosylphosphatidylinositol-anchored protein located on the plasma membrane. In this study, we used CRISPR/Cas9 to target the second and third exons of theSRL1gene in the indica rice line GXU103, which resulted in the generation of 14 T0transgenic plants with a double-target mutation rate of 21.4%. After screening 120 T1generation plants, we identified 26 T-DNA-free homozygous double-target mutation plants. We designated the resultingSRL1homozygous double-target knockout assrl1-103. This line exhibited defects in leaf development, leaf rolling in the mature upright leaves, and a compact nature of the fully grown plants. Compared with the wild type (WT), the T2generation ofsrl1-103varied in two key aspects: the width of flag leaf (12.6% reduction compared with WT) and the leaf rolling index (48.77% increase compared with WT). In order to gain a deeper understanding of the involvement ofSRL1in the regulatory network associated with rice leaf development, we performed a transcriptome analysis for the T2generation ofsrl1-103. A comparison ofsrl1-103with WT revealed 459 differentially expressed genes (DEGs), including 388 upregulated genes and 71 downregulated genes. In terms of the function of the DEGs, there seemed to be a significant enrichment of genes associated with cell wall synthesis (LOC_Os08g01670,LOC_Os05g46510,LOC_Os04g51450,LOC_Os10g28080,LOC_Os04g39814,LOC_Os01g71474,LOC_Os01g71350, andLOC_Os11g47600) and vacuole-related genes (LOC_Os09g23300), which may partially explain the increased leaf rolling insrl1-103. Furthermore, the significant downregulation of BAHD acyltransferase-like protein gene (LOC_Os08g44840) could be the main reason for the decreased leaf angle and the compact nature of the mutant plants. In summary, this study successfully elucidated the gene regulatory network in whichSRL1participates, providing theoretical support for targeting this gene in rice breeding programs to promote variety improvement.