Protocols used in RNA Extraction ================================ Long Cytoplasmic RNA A+ and A-: Long Cytoplasmic RNA was isolated using Qiagens RNeasy Protocol. Qiagens Qligotex kit was used to separate Poly-A(+) from Poly-A(-). Long Nuclear RNA A+ and A-: Cell were lysed with Qiagens RLN buffer and the nuclei were spun out, resuspended in Qiagen's RLT buffer, layered over a Cesium Chloride bed and spun at 24,000 rpm for a minimum of 20 hours. The pellet was recovered and cleaned up on Qiagens RNeasy kit. Qiagens Qligotex kit was used to separate Poly-A(+) from Poly-A(-). Long Polysomal RNA A+ and A-: Cyclohexamide arrested cell lysate were layered on a 20-60% sucrose gradient and spun at 27,000 rpm for 4 hours. The fractions and analyzed by UV-spectroscopy using a programmable Density Gradient Fractionation System Foxy Jr. (Isco). Long Nucleoplasm, Nucleoli and Chromatin: We used the protocol found in Bhorjee and Pederson (1973). Small Cytoplasmic RNA: Small Cytoplasmic RNA was isolated using Qiagens RNA/DNA kit. Small Nuclear RNA: Small Nuclear RNA was isolated using Qiagens RNA/DNA kit. References ========== Bhorjee JS, Pederson T. Chromatin: its isolation from cultured mammalian cells with particular reference to contamination by nuclear ribnucleoprotein particles. Biochemistry. 1973 Jul 3;12(14):2766-73. Kapranov, P. et.al. RNA maps reveal new RNA classes and a possible function for pervasive transcription. Science. 2007 Jun 8;316(5830):1484-8.