Components

10X Glycine10X PBSNuclei Isolation BufferRIP Cross-Linked Lysis BufferProtein A/G Magnetic BeadsNuclear RIP Dilution BufferLow Salt Wash BufferHigh Salt Wash BufferLiCl Wash BufferTE BufferRIP Elution Buffer10% SDS0.5 M EDTADNase I (RNase Free)DNase I SupplementDNase I Reaction Buffer Protease Inhibitor Cocktail III, Animal Free RNAse InhibitorProteinase K
Control Antibodies and Primers
Normal Mouse IgG Negative Control AntibodyAnti-EZH2 Positive Control Antibody NEAT1 Positive Control PrimersU1 snRNA Negative Control Primers

General description

Features and Advantages Generates cross-linked chromatin to allow analysis of a variety of chromatin-associated RNAs Flexible, scalable input requirements: Recover RNA from millions of cells or as few as 5,000 cells Magnetic protein A/G bead blend and optimized buffer system results in low backgrounds and high signal-to-noise ratios Suitable for analysis by RT-qPCR or RIP-seq Complete set of reagents and detailed protocol to enable first-time success Magna Nuclear RIP™ Kits are specially designed to allow the discovery and analysis of a variety of chromatin associated RNAs such as long non-coding RNAs, enhancer RNAs and miRNAs . These chromatin-associated RNAs often regulate gene expression and can be analyzed with applications including quantitative reverse transcription polymerase chain reaction (RT-PCR), microarray analysis (RIP-chip) and next generation sequencing (RIP-Seq).Nuclear RIP can be performed using chromatin that has interactions stabilized by formaldehyde treatment (cross-linked) or chromatin that has not been treated with a cross linking reagent (native). While both of these approaches are similar in that they are designed to recover chromatin associated RNA, the reagents used and the details of the protocol and types of interactions typically detected are different. Cross-linked can capture higher molecular weight complexes in in vivo configurations with possibly lower affinities. In contrast native RIP is expected to recover high affinity, more direct interactions between proteins encoded RNA binding motifs and candidate RNAs. For less well understood proteins and protein complexes often both approaches are used. The kit described here is for the cross-linked approach. If a native approach is of interest please visit the product page for the Magna Nuclear RIP (Native) Kit, catalogue # 17-10522 or the EZ-Magna Nuclear RIP (Native) Kit, catalogue # 17-10523.

Legal Information

NuCLEAR is a trademark of Sigma-Aldrich Co. LLC

Packaging

Kit capacity: 24 RNA-binding protein immunoprecipitation assays using a cross-linked nuclear lysate

Physical form

Two boxes containing key reagents for generation of cross linked nuclear lysates and performance of 24 individual RNA-binding protein immunoprecipitation (RIP) reactions. Plus box containing positive and negative control antibodies and qPCR primers.

Storage and Stability

Upon receipt, store components at the temperatures indicated on the labels.
Kit components are stable for 6 months from date of shipment when stored as directed.