|Description||EZ- Magna ChIRP RNA Interactome Kit -Isolation and characterization of non-coding RNA:chromatin complexes|
|Overview||Features and Advantages
The Magna ChIRP RNA Interactome Kit uses the ChIRP method (Chromatin Isolation by RNA Purification) to enable identification and analysis of regions of chromatin that interact with a particular chromatin-associated RNA. This novel anti-sense probe-based capture strategy uses cross-linked chromatin to allow unbiased discovery of RNA-associated DNA sequences and proteins. To perform ChIRP two pools consisting of mulitple biotinylated oligonucleotide probes complementary to the RNA of interest are used (see figure 1). These probe sets are combined with chromatin and hybridized to the chromatin-associated RNA. Complexes containing biotinylated-probes bound to the chromatin-associated RNA are then isolated using streptavidin magnetic beads. DNA or RNA can then be recovered and analyzed by quantitative PCR or next generation sequencing (ChIRP-seq). Alternatively, proteomic analysis of the recovered chromatin can be performed.
Learn more about the ChIRP method in JOVE! (Watch Now!)
Need help making your own ChIRP probe sets? (Use the ChIRP probe designer tool.)
|Materials Required but Not Delivered||Reagents
|Presentation||Three boxes containing key reagents for the extraction of ChIRP ready chromatin, plus magnetic beads, enzymes, buffers, biotinlyated negative control probes, biotinylated TERC odd and even positive control probe set and PCR primers to allow the performance of 12 chromatin isolation by RNA purification assays.|
|Safety Information according to GHS|
|Storage and Shipping Information|
|Storage Conditions||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.
|Material Size||12 Assays|
|Material Package||Kit capacity: 12 Chromatin Isolation by RNA Purification Assays (includes negative control probes, even and odd positive control TERC lncRNA probes, and control PCR primer sets)|
EZ- Magna ChIRP RNA Interactome Kit -Isolation and characterization of non-coding RNA:chromatin complexes SDS
|Reference overview||Pub Med ID|
|Long noncoding RNA BCAR4 promotes osteosarcoma progression through activating GLI2-dependent gene transcription.|
Chen, F; Mo, J; Zhang, L
Tumour Biol 13403-13412 2016
Despite great advances have been made in the understanding of biology of osteosarcoma, the molecular mechanisms involved in osteosarcoma tumorigenesis and progression are still largely unknown. Long noncoding RNA (lncRNA) is a new type of RNA molecule, which plays pivotal roles in many tumors. lncRNA BCAR4 has been identified as an oncogenetic lncRNA involved in the progression of breast cancer. However, the functions and clinical significances of BCAR4 in osteosarcoma are unknown now. In this study, we found that BCAR4 was significantly upregulated in osteosarcoma tissues. Increased expression of BCAR4 was significantly correlated with large tumor size, advanced Enneking stage, lung metastasis, and poor prognosis. Functional experiments demonstrated that knockdown of BCAR4 inhibits the proliferation and migration of osteosarcoma cell in vitro. Consistently, knockdown of BCAR4 inhibits osteosarcoma tumorigenesis and lung metastasis in vivo. Chromatin isolation by RNA purification assay showed that BCAR4 physically associates with the promoters of GLI2 target genes. The depletion of BCAR4 inhibits the expression of GLI2 target genes and GLI2 reporter luciferase activity in a dose-dependent manner. The expression of BCAR4 and GLI2 target genes is significantly correlated in osteosarcoma tissues. Depletion of DLI2 abolished the effects of BCAR4 on osteosarcoma. Taken together, these findings demonstrated that BCAR4 promotes osteosarcoma progression via activating GLI2-dependent gene transcription and serves as a potential prognostic biomarker and a therapeutic target of osteosarcoma.
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Instructions for Use
|EZ- Magna ChIRP RNA Interactome Kit Analysis and Mapping of Chromatin: Noncoding RNA Interactions|