RNeasy Plus Mini Kit

細胞および組織から100 µgまでのトータルRNA精製用(gDNA Eliminatorカラム付き)

  • ユニークなgDNA EliminatorカラムによりDNase 分解不要
  • ゲノムDNAの効率的な除去
  • 短時間で再現性の高いRNA
  • 高感度アプリケーションで良好な結果を実現するRNA
RNeasy Plus Mini Kitは100 µg RNAまでの迅速で便利な精製と効率的なゲノムDNA除去を組み合わせています。まず細胞あるいは組織ライセートをgDNA Eliminator Spin Columnに通過させて、ゲノムDNAを除去します。続いてRNeasy Mini Spin Columnを用いてトータルRNAを精製します。本キットはQIAcube上で自動化可能です。組織サンプルはRNAlater RNA Stabilization ReagentまたはAllprotect Tissue Reagentで簡便に安定化され、TissueRuptorあるいはTissueLyserシステムで効率的に破砕されます。少量のサンプルにはRNeasy Plus Micro Kit(スピンカラム結合容量は 45 µg RNA)をご使用ください。
Product Cat. no. List price:
RNeasy Plus Mini Kit (50)
For 50 minipreps: RNeasy Mini Spin Columns, gDNA Eliminator Spin Columns, Collection Tubes, RNase-Free Water and Buffers
74134
HK$3,290.00
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RNeasy Plus Mini Kit (250)
For 250 minipreps: RNeasy Mini Spin Columns, gDNA Eliminator Spin Columns, Collection Tubes, RNase-Free Water and Buffers
74136
HK$14,460.00
Add to cart
The RNeasy Plus Mini Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.
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High, reproducible RNA yields.|Effective tissue genomic DNA removal.|RNeasy Plus procedure.|Effective cell genomic DNA removal.|Array-ready RNA.|
Total RNA was purified in duplicate from different amounts of Jurkat cells using the RNeasy Plus Mini Kit or a similar kit from Supplier AV. Real-time RT-PCR assays for β-actin were performed (40 cycles). The lower CT values with the RNeasy Kit demonstrate greater RNA yields. With the kit from Supplier AV, no transcript was detectable in RNA purified from 102 cells. |Total RNA was purified in duplicate from various mouse tissues (10 mg per sample) using the RNeasy Plus Mini Kit or kits from other suppliers. Real-time PCR assays for c-jun were performed to determine the amount of DNA contamination in the purified RNA.|A short workflow enables RNA purification with genomic DNA removal in less than 25 minutes.|Total RNA was purified from Jurkat cell samples (1 x 106 cells per sample) using [A] the RNeasy Plus Mini Kit, or [B] an RNA purification kit with integrated genomic DNA removal from Supplier AV. Duplicate real-time RT-PCR assays for β-actin were performed with (+RT) or without (-RT) reverse transcriptase. The -RT curves demonstrate that RNA purified using the RNeasy Plus Mini Kit was virtually free of genomic DNA.|Total RNA was purified in duplicate from 1 x 106 HeLa cells using the RNeasy Plus Mini Kit. cRNA was prepared from the duplicate RNA samples (3.5 μg each) using the GeneChip IVT Labeling Kit. The cRNA samples (15 μg each) were analyzed on GeneChip Human Genome U133A probe arrays. The scatter plot shows the correlation between the two samples (Pearson correlation coefficient [r] is 0.996). Red: gene present in both samples; Blue: gene absent or marginal in one sample; Yellow: gene absent or marginal in both samples.|
Performance
RNeasy Plus Mini Kitによる細胞および組織からのRNA精製では、ゲノムDNAの効率的な排除と、再現性および収量の高いRNA精製を実現し、精製したRNAは感度の高いアプリケーションに使用できます(図"効率的な細胞ゲノムDNAの除去"、"効率的な組織ゲノムDNAの除去"、"再現性の高いRNA収量"、"アレイ解析が即可能なRNA")。Agilent RIN 値が約10 のトータルRNA が培養細胞からのルーチン精製で得られます。
Principle

まず細胞を高度変性用グアニジンイソチオシアネート含有のBuffer RLT Plus中で溶解・ホモジナイズし、即座にRNase を不活化することで、劣化のないRNAを確実に分離します。次に、ライセートをgDNA Eliminator Spin columnに通し、高塩濃度バッファーを併用することでゲノムDNAを迅速かつ効率的に除去します。エタノールを添加してRNAの結合条件を最適にし、サンプルをRNeasy MinElute spin columnにアプライします。このRNA精製専用カラムには、溶解細胞からのRNA を特異的に結合するシリカメンブレンが充填されています。

Procedure

RNeasy Plus Mini Kitは107個までの細胞または30 mgまでの組織からトータルRNAを精製します。簡単な操作手順によりゲノムDNA除去を含むRNA精製が25分未満で行なえます(フローチャート"RNeasy Plus操作手順")。RNA の分離にはまずサンプルを溶解し、ホモジナイズします。ライセートをgDNA Eliminator spin columnに通し、ろ液にエタノールを加えRNeasy MinElute spin columnにアプライします。RNA はメンブレンに結合し、夾雑物は洗い流されます。高品質RNAが30 µl以上の水で溶出されます。

本キットでは、様々なスタートサンプルに最適化されたプロトコールを用意しています。それぞれのプロトコールは、主にサンプルの溶解およびホモジナイズの方法が異なり、サンプルをgDNA Eliminator spin columnにアプライした後のプロトコールはほとんど共通しています。本操作ではrRNAsやtRNAsなどの200ヌクレオチド未満のRNAはほとんど除かれるため、mRNAリッチなRNAを精製できます。培養細胞からmiRNAのような低分子RNAを含むトータルRNAを精製したい場合は、RNeasy PlusKitの操作手順を変更することができます。 

Buffer RLT Plus(RNeasy Plus Mini Kitに添付)で組織を破砕およびホモジナイズする時、気泡が大量に生じることがあります。破砕とホモジナイゼーションの前に、Buffer RLT PlusにReagent DX(別売)を添加すると、気泡の発生が大幅に抑えられます。Reagent DXは本キットで十分にテストされており、RNA純度あるいはダウンストリームアプリケーションへの影響はありません。

Applications

RNeasy Plus Mini Kitを用いて精製したRNAは、リアルタイム定量RT-PCRのような微量のDNAコンタミにも敏感なダウンストリームアプリケーションにも最適です。精製されたRNAは、その他のアプリケーションにも利用可能です。

Feature
Specifications
Applications Northern, dot, and slot blotting, end-point RT-PCR, quantitative, real-time RT-PCR, array analysis, next-generation sequencing
Elution volume 30-50 µl
Format Spin column
Integrated removal of genomic DNA Yes
Main sample type Tissue, cells
Processing Manual (centrifugation)
Purification of total RNA, miRNA, poly A+ mRNA, DNA or protein Total RNA
Sample amount 30 mg (<700 µl)
Technology Silica technology
Time per run or per prep 25 minutes
Yield Varies

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Supplementary Protocols
2

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There are two protocols: follow Protocol 1 if you want to purify total RNA containing miRNA, or follow Protocol 2 if you want to purify small RNA (includes miRNA, 5S rRNA, and tRNA) and larger RNA (>200 nt) separately.
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Safety Data Sheets
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Scientific Posters
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Images
High, reproducible RNA yields.
High, reproducible RNA yields.
Total RNA was purified in duplicate from different amounts of Jurkat cells using the RNeasy Plus Mini Kit or a similar kit from Supplier AV. Real-time RT-PCR assays for β-actin were performed (40 cycles). The lower CT values with the RNeasy Kit demonstrate greater RNA yields. With the kit from Supplier AV, no transcript was detectable in RNA purified from 102 cells. 
Effective genomic DNA removal.
Effective tissue genomic DNA removal.

Total RNA was purified in duplicate from various mouse tissues (10 mg per sample) using the RNeasy Plus Mini Kit or kits from other suppliers. Real-time PCR assays for c-jun were performed to determine the amount of DNA contamination in the purified RNA.

RNeasy Plus procedure.
RNeasy Plus procedure.

A short workflow enables RNA purification with genomic DNA removal in less than 25 minutes.

Effective elimination of genomic DNA contamination.
Effective cell genomic DNA removal.
Total RNA was purified from Jurkat cell samples (1 x 106 cells per sample) using [A] the RNeasy Plus Mini Kit, or [B] an RNA purification kit with integrated genomic DNA removal from Supplier AV. Duplicate real-time RT-PCR assays for β-actin were performed with (+RT) or without (-RT) reverse transcriptase. The -RT curves demonstrate that RNA purified using the RNeasy Plus Mini Kit was virtually free of genomic DNA.
Array-ready RNA.
Array-ready RNA.
Total RNA was purified in duplicate from 1 x 106 HeLa cells using the RNeasy Plus Mini Kit. cRNA was prepared from the duplicate RNA samples (3.5 μg each) using the GeneChip IVT Labeling Kit. The cRNA samples (15 μg each) were analyzed on GeneChip Human Genome U133A probe arrays. The scatter plot shows the correlation between the two samples (Pearson correlation coefficient [r] is 0.996). Red: gene present in both samples; Blue: gene absent or marginal in one sample; Yellow: gene absent or marginal in both samples.