HT-29 cells were transfected with miR-17-5p mimic or miR-19-3p. After 48 h, the cells were lysised in NP40 buffer supplemented with RNase inhibitor, protease inhibitor and phosphatase inhibitor (Sigma). The samples were then centrifuged for 15 min at 12,000 g at 4°C in a microcentrifuge. The supernatant was placed in a fresh tube kept on ice and subjected to Ago2 immunoprecipitation using the miRNA isolation kit for human Ago2 (Wako, Japan) according to the manufacturer recommendation. RNA derived from transfected cells following Ago2 immunoprecipitation were analyzed by real-time PCR.
Mirna isolation kit for human ago2
Manufactured by Fujifilm
Sourced in Japan
The MiRNA isolation kit for human Ago2 is a laboratory equipment product designed to isolate microRNA (miRNA) that is bound to the Argonaute 2 (Ago2) protein. The kit provides the necessary reagents and protocols to efficiently extract and purify miRNA-Ago2 complexes from biological samples.
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Lab products found in correlation
2 protocols using mirna isolation kit for human ago2
1
Profiling miRNA-Protein Interactions
2
Quantification of miRNAs Bound to Ago2
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To explore whether both exogenous strands of pre‐miR‐455 (miR‐455‐5p and miR‐455‐3p) were incorporated into the RNA‐induced silencing complex (RISC), we performed Ago2 immunoprecipitation assays using a miRNA isolation kit for human Ago2 (Wako, Osaka, Japan). The procedure was described previously (Goto et al., 2017 ; Okato et al., 2017a ). Quantification of miRNAs bound to Ago2 was achieved by qRT‐PCR, and normalization was performed relative to the expression of miR‐26a that is unaffected by transfection with miR‐455‐5p/‐3p. Details of the reagents used are shown in Table S2 .
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