Splicing plays a major role in eukaryotic gene expression by processing pre-mRNA to form mature mRNA. Splicing is catalysed by the spliceosome; a ribonucleoprotein complex consisting of five small nuclear ribonucleoproteins (snRNPs) and over 100 proteins. Proper assembly of spliceosomal components is critical for its function, and thus assembly defects can be lethal. Several diseases have been associated with splicing defects, such as cancer (breast cancer, leukaemia), cystic fibrosis, duchenne muscular dystrophy (DMD), retinitis pigmentosa (RP) and taybi-linder syndrome (TALS). Studying the structural dynamics and distinct functions of snRNA complexes and the factors that affect the stability of those complexes provides an overall idea regarding the structure and function of the spliceosome, which can guide us to discover novel therapeutics for splicing related diseases. Hence, the aim of this study is employing smFRET technique to monitor the structural dynamics and assembly of snRNA complexes and the effect of protein factors on those dynamics with single molecule resolution. Three specific aims have been addressed in this thesis work to achieve the main goal. The first part of this study is focused on understanding how spliceosomal components are recycled. This work shows that binding of Prp24; U6 snRNP specific protein unwinds U2 from U2/U6 complex and stabilizes U6 at a low FRET conformation, suggesting a novel role for Prp24 as a recycling factor. The second part of this study is focused on understanding the assembly of sub-spliceosomal complexes and their global structure. This work shows that although the binding of individual proteins slightly changes the conformation of U4/U6 duplex; overall it maintains a rigid structure. This suggests that the U4/U6 adopts a preformed conformation and act as a scaffold for protein binding, while preventing U6 from premature activation. The third part of this study is focused on understanding structural and functional similarities between minor and major spliceosomal complexes. This work shows that minor spliceosomal U12/U6atac complex adopts a conformation similar to the three-helix junction structure of major spliceosomal U2/U6.