کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
10826607 | 1064804 | 2005 | 11 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
CLIP: A method for identifying protein-RNA interaction sites in living cells
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موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
زیست شیمی
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چکیده انگلیسی
Nucleic-acid binding proteins constitute nearly one-fourth of all functionally annotated human genes. Genome-wide analysis of protein-nucleic acid contacts has not yet been performed for most of these proteins, restricting attempts to establish a comprehensive understanding of protein function. UV cross-linking is a method typically used to determine the position of direct interactions between proteins and nucleic acids. We have developed the cross-linking and immunoprecipitation assay, which exploits the covalent protein-nucleic acid cross-linking to stringently purify a specific protein-RNA complex using immunoprecipitation followed by SDS-PAGE separation. In this way, the vast majority of non-specific contaminating RNA, which can bind to co-immunoprecipitated proteins or beads, can be removed. Here, we present an improved protocol that performs RNA linker ligation before the SDS-PAGE step, and describe its application to the specific purification and amplification of RNA ligands of Nova in neurons.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Methods - Volume 37, Issue 4, December 2005, Pages 376-386
Journal: Methods - Volume 37, Issue 4, December 2005, Pages 376-386
نویسندگان
Jernej Ule, Kirk Jensen, Aldo Mele, Robert B. Darnell,