Alternative splicing (AS) significantly enhances transcriptome and proteome diversity, playing a critical role in eukaryotic physiology and development. With advances in high-throughput sequencing, many novel transcript isoforms and splicing-related factors have been identified. This review summarizes splicing mechanisms in prokaryotes and eukaryotes, recent studies on AS in diverse plant species and under various abiotic stresses, and modern techniques for studying AS functions and protein products. By integrating genetic studies, quantitative methods, and high-throughput omics, researchers can discover novel transcript isoforms and functional splicing factors, enhancing understanding of splicing modes in plants. AS is essential for gene expression, influencing development, stress response, and DNA damage repair in eukaryotes. In plants, up to 90% of genes contain introns, and about 42-61% undergo AS. AS contributes more to mRNA transcript diversity than proteome diversity. AS regulates protein expression by modifying transcripts, influencing isoform stability or ratios. AS is crucial for plant development, stress responses, and phenotypic traits. In animals, AS plays roles in various biological processes and can cause disease if uncontrolled. AS increases genetic instability, linked to tumorigenesis. In plants, AS is involved in various developmental processes and abiotic stress responses, including circadian clock, flowering, flooding, and drought. Splicing factors have been identified through bioinformatics and gene verification, influencing splicing core components and pathways. For example, the A.thaliana atu2af65b mutant reduces FLC transcript abundance, affecting flowering. Light-activated photoreceptors regulate AS in Physcomitrella patens to modulate phototropic responses. Splicing mechanisms in eukaryotic cells involve the spliceosome, which catalyzes intron removal and exon joining. Two types of spliceosomes, the major U2-dependent and U12-dependent, are involved. Splice sites are recognized by snRNPs, and splicing involves two steps: intron removal and exon joining. Spliceosome assembly and function are crucial for accurate splicing. Prokaryotic splicing is less common due to polycistronic genes and coupled transcription-translation. Prokaryotes use group I and II introns, which self-splice. Eukaryotic splicing is more complex, involving multiple steps and various splicing types. Eukaryotic splicing machinery includes snRNPs and proteins, while prokaryotic splicing relies on self-splicing introns. In plants, AS is crucial for development and stress responses. AS events include IR, ES, and ME. AS regulates gene expression during development, affecting processes like circadian clock, root and embryo development, flowering, and fruit ripening. AS is also involved in stress responses, such as drought, salt, and cold stress. Splicing factors and spliceosome components regulate AS, influencing plantAlternative splicing (AS) significantly enhances transcriptome and proteome diversity, playing a critical role in eukaryotic physiology and development. With advances in high-throughput sequencing, many novel transcript isoforms and splicing-related factors have been identified. This review summarizes splicing mechanisms in prokaryotes and eukaryotes, recent studies on AS in diverse plant species and under various abiotic stresses, and modern techniques for studying AS functions and protein products. By integrating genetic studies, quantitative methods, and high-throughput omics, researchers can discover novel transcript isoforms and functional splicing factors, enhancing understanding of splicing modes in plants. AS is essential for gene expression, influencing development, stress response, and DNA damage repair in eukaryotes. In plants, up to 90% of genes contain introns, and about 42-61% undergo AS. AS contributes more to mRNA transcript diversity than proteome diversity. AS regulates protein expression by modifying transcripts, influencing isoform stability or ratios. AS is crucial for plant development, stress responses, and phenotypic traits. In animals, AS plays roles in various biological processes and can cause disease if uncontrolled. AS increases genetic instability, linked to tumorigenesis. In plants, AS is involved in various developmental processes and abiotic stress responses, including circadian clock, flowering, flooding, and drought. Splicing factors have been identified through bioinformatics and gene verification, influencing splicing core components and pathways. For example, the A.thaliana atu2af65b mutant reduces FLC transcript abundance, affecting flowering. Light-activated photoreceptors regulate AS in Physcomitrella patens to modulate phototropic responses. Splicing mechanisms in eukaryotic cells involve the spliceosome, which catalyzes intron removal and exon joining. Two types of spliceosomes, the major U2-dependent and U12-dependent, are involved. Splice sites are recognized by snRNPs, and splicing involves two steps: intron removal and exon joining. Spliceosome assembly and function are crucial for accurate splicing. Prokaryotic splicing is less common due to polycistronic genes and coupled transcription-translation. Prokaryotes use group I and II introns, which self-splice. Eukaryotic splicing is more complex, involving multiple steps and various splicing types. Eukaryotic splicing machinery includes snRNPs and proteins, while prokaryotic splicing relies on self-splicing introns. In plants, AS is crucial for development and stress responses. AS events include IR, ES, and ME. AS regulates gene expression during development, affecting processes like circadian clock, root and embryo development, flowering, and fruit ripening. AS is also involved in stress responses, such as drought, salt, and cold stress. Splicing factors and spliceosome components regulate AS, influencing plant