RNA-Seq of crRNAs bound by Sa. islandicus type I-A Cascade complex
The crRNA- Para-csa5His were obtained from the typeI-A CRISPR Cascade complex which were expressed in Saccharolobus islandicus REY15A cells . The archaeal cells were lysed, and the typeI-A Cascade complex was purified via affinity chromatography (using a His-tag). The crRNA was subsequently extracted from the purified complex. Then crRNAs were processed with T4 polynucleotide kinase (T4 PNK) to generate 5monophosphate ends suitable for small RNA library. Small RNA library preparation and sequencing using the NEXTflex Small RNA-Seq Kit v3 from Bioo Scientific.