Kit for simply and conveniently detecting RNA and protein interaction and usage method thereof

A protein interaction and kit technology, used in biological testing, material testing, etc., can solve the problem of lack of RNA and protein, and achieve the effect of short time-consuming, simple operation and wide application.

Active Publication Date: 2016-11-09
广州赛诚生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In summary, there is no particularly robust and time-saving method in the art for detecting RNA-protein interactions

Method used

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  • Kit for simply and conveniently detecting RNA and protein interaction and usage method thereof
  • Kit for simply and conveniently detecting RNA and protein interaction and usage method thereof
  • Kit for simply and conveniently detecting RNA and protein interaction and usage method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0058] A simple kit for detecting RNA-protein interaction, including biotin-labeled UTP, ATP, CTP, GTP mixture, T7 RNA polymerase, cell lysate, streptavidin magnetic beads, washing solution and protein eluent .

[0059] Wherein, the formula of the cell lysate includes the following raw materials:

[0060] 1% NP-40;

[0061] 0.25% sodium deoxycholate;

[0062] Tris-HCL 50mmoL / L;

[0063] Nacl 150mmoL / L;

[0064] EDTA 1mmoL / L;

[0065] PMSF 1mmoL / L;

[0066] The pH value of the Tris-HCL is 7.4.

[0067] The formula of described lotion comprises following raw material:

[0068] Tris-HCL 50mmoL / L;

[0069] EDTA 1mM;

[0070] KCl 100Mm;

[0071] Triton X-100;

[0072] 0.1% glycerol;

[0073] 5%DTT 1mM;

[0074] The pH value of the Tris-HCL is 7.0.

[0075] The formula of described protein eluent comprises following raw material:

[0076] 250 mM Tris-HCl;

[0077] 10% (W / V) SDS;

[0078] 0.5% (W / V) bromophenol blue;

[0079] 50% (V / V) glycerin;

[0080] 5% (W / V) β...

Embodiment 2

[0097] A simple kit for detecting RNA-protein interaction, including biotin-labeled UTP, ATP, CTP, GTP mixture, T7 RNA polymerase, cell lysate, streptavidin magnetic beads, washing solution and protein eluent .

[0098] Wherein, the formulation of the cell lysate includes the following raw materials:

[0099] 0.05% by volume NP-40;

[0100] 0.15% by volume sodium deoxycholate;

[0101] Tris-HCL 45mmoL / L;

[0102] Nacl 140mmoL / L;

[0103] EDTA 0.8mmol / L;

[0104] PMSF 0.8mmoL / L;

[0105] The pH value of the Tris-HCL is 7.2.

[0106] The formula of described lotion comprises following raw material:

[0107] Tris-HCL 45mmoL / L;

[0108] EDTA0.8mM;

[0109] KCl 80mM;

[0110] 0.1% by volume TritonX-100;

[0111] 0.05% by volume glycerol;

[0112] 4.5% by volume DTT;

[0113] The pH value of the Tris-HCL is 6.8.

[0114] The formula of described protein eluent comprises following raw material:

[0115] 220 mM Tris-HCl;

[0116] 8.5% (W / V) SDS;

[0117] 0.4% (W / V) br...

Embodiment 3

[0129] A simple kit for detecting RNA-protein interaction, including biotin-labeled UTP, ATP, CTP, GTP mixture, T7 RNA polymerase, cell lysate, streptavidin magnetic beads, washing solution and protein eluent .

[0130] Wherein, the formula of the cell lysate includes the following raw materials:

[0131] 1.5 volume% NP-40;

[0132] 0.35% by volume sodium deoxycholate;

[0133] Tris-HCL 55mmoL / L;

[0134] Nacl 160mmoL / L;

[0135] EDTA 1.2mmol / L;

[0136] PMSF 1.2mmoL / L;

[0137] The pH value of the Tris-HCL is 7.5.

[0138] The formula of described lotion comprises following raw material:

[0139] Tris-HCL 55mmoL / L;

[0140] EDTA 1.2 mM;

[0141] KCl 120mM;

[0142] 0.2% by volume TritonX-100;

[0143] 0.15% by volume glycerol;

[0144] 5.5% by volume DTT;

[0145] The pH value of the Tris-HCL is 7.2.

[0146] The formula of described protein eluent comprises following raw material:

[0147] 280 mM Tris-HCl;

[0148] 11.5% (W / V) SDS;

[0149] 0.6% (W / V) bromop...

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Abstract

The invention discloses a kit for simply and conveniently detecting RNA and protein interaction and a usage method thereof. The kit contains a UTP, ATP, CTP and GTP mixture marked with biotin, T7RNA polymerase, a cell lysis solution, streptavidin magnetic beads, lotion and protein eluent. The kit utilizes RNA to catch protein producing interaction with the RNA, RNA (such as biotin) with known sequences are firstly marked, are fixed to Beads by relying on the marks and then are co-incubated with a substance containing protein, then supernatant which does not react with the RNA is removed, the protein washed from the Beads is collected, and the obtained protein can be used for mass spectrometry or western blot experiment.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to a kit for conveniently detecting the interaction between RNA and protein and its application method. Background technique [0002] Non-coding RNAs are key regulators of many cellular processes, are critical to maintaining cellular homeostasis, and are closely related to development and cancer progression. Noncoding RNAs mainly include long noncoding RNAs (lncRNAs) and microRNAs (miRNAs), which can regulate the expression of transcription factors, guide chromatin reorganization and modification, reduce mRNA translation and control gene expression. [0003] RNA-binding proteins (RNA-binding proteins) play an important role in the regulation of post-transcriptional gene expression, and they regulate cell functions by interacting with RNA. RNA-binding proteins are involved in various aspects of RNA metabolism, including RNA splicing, polyadenylation, sequence editing, RNA transport,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68
CPCG01N33/68
Inventor 高飞钱智鹏乔继彪张映菲张余琴
Owner 广州赛诚生物科技有限公司
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