FastLane Cell cDNA Kit

For high-speed preparation of cDNA without RNA purification for use in real-time RT-PCR
  • From cells to cDNA in only 4 steps and under 45 minutes
  • Sensitive detection of even low-abundance transcripts
  • Easy parallel processing of several samples
  • Integrated gDNA removal allows detection of RNA only
  • No need to design RNA-specific primers or probes

The FastLane Cell cDNA Kit provides a fast and simple procedure for preparing first-strand cDNA directly from cultured cells without RNA purification. The kit is supplied with wash and lysis buffers for preparing lysates and stabilizing RNA, gDNA Wipeout Buffer for eliminating genomic DNA contamination, and all reaction components for fast and efficient cDNA synthesis. The FastLane Cell cDNA Kit is ideal for experiments requiring rapid, high-throughput gene expression analysis.

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製品名 Cat. no. List price:
FastLane Cell cDNA Kit (50)
Buffer FCW, Buffer FCP, and components for 50 x 20 µl reverse-transcription reactions (gDNA Wipeout Buffer, Quantiscript Reverse Transcriptase, Quantiscript RT Buffer, RT Primer Mix, and RNase-Free Water)
215011
¥74,000

FastLane Cell cDNA Kit  は分子生物学実験用です。疾病の診断、治療または予防の目的には使用することはできません。


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がん研究において最高の感度を実現|CDC2ノックダウンの優れた解析結果|ゲノムDNAコンタミを効率的に除去|再現性の高いcDNA調製|
HeLa細胞を用いたがん研究に重要な様々な遺伝子の2ステップリアルタイムRT-PCR解析において、Supplier AIVの類似キットの代わりにFastLane Kitを用いてcDNAを調製した時に、より低いCT値が得られた。| 様々なCDC2 siRNAs(S1S2S3)あるいはnonsilencing siRNA(NS)をMCF-7細胞にトランスフェクトした。48時間後にFastLane Kitを用いてcDNAを調製した。GAPDH(内因性コントロール)に対するCDC2(細胞周期関連タンパク質)の相対的な発現を2ステップリアルタイムRT-PCRにより解析した。未処理細胞中(UT)の相対的なCDC2発現を100%とした。 |FastLane Cell cDNA Kitを用いて逆転写酵素を添加(+RT)あるいは未添加(-RT)でcDNAを調製した。β-actinの遺伝子発現アッセイを用いてcDNAを定量した。–RTの線は、ゲノムDNAが検出されなかったことを示唆している。|HeLa細胞を含む48ウェルプレートからFastLane Kitを用いてcDNAを調製した。Tubulin B(構造タンパク質)の発現を2ステップリアルタイムRT-PCRにより解析した。|
パフォーマンス
cDNA synthesized by the FastLane Cell cDNA Kit enables highly sensitive and reproducible results in real-time two-step RT-PCR (see figures "Superior sensitivity in cancer research" and "Highly reproducible cDNA preparation"). The high quality of the cDNA also allows detection over a wide linear range from >105 cells down to one cell.
原理

The FastLane Cell cDNA Kit uses FastLane technology to prepare cDNA directly from cultured cells in just 45 minutes. To ensure only RNA is detected in real-time RT-PCR, novel gDNA Wipeout Buffer is used to eliminate genomic DNA (see figure "Effective elimination of genomic DNA contamination"). Time and effort are saved as there is no need to design RNA-specific primers or probes.

When the FastLane Cell cDNA Kit is used together with optimized QuantiTect, QuantiFast, or Rotor-Gene Kits for real-time RT-PCR, several cell samples can be easily processed and analyzed within a few hours. This allows a snapshot of several transcripts in experiments such as validation of siRNA-mediated gene knockdown (see figure "Successful analysis of CDC2 knockdown").

The FastLane Cell cDNA Kit is part of the comprehensive range of FastLane Kits, which speed up and streamline the gene expression analysis workflow.

操作手順

With the FastLane Cell cDNA Kit, cDNA is obtained from cultured cells in just 45 minutes in a 4-step procedure: cell wash; cell lysis with integrated RNA stabilization; genomic DNA elimination; and cDNA synthesis. The synthesized cDNA is ready to use in real-time two-step RT-PCR.

The kit contains sufficient reagents for preparing cDNA from two 24-well cell-culture plates or one 48-well cell-culture plate. If working with a 96-well cell-culture plate, an additional QuantiTect Reverse Transcription Kit (50) needs to be purchased.

cDNA prepared with the FastLane Cell cDNA Kit can be analyzed by real-time two-step RT-PCR using one of the following QuantiTect Kits:

  • SYBR® Green detection: QuantiTect SYBR® Green PCR Kit
  • Probe detection: QuantiTect Probe PCR Kit
  • Multiplex probe detection: QuantiTect Multiplex PCR Kits

Alternatively, fast real-time two-step RT-PCR can be carried out when the FastLane Cell cDNA Kit is used in combination with a QuantiFast Kit:

  • SYBR® Green detection: QuantiFast SYBR® Green PCR Kit
  • Probe detection: QuantiFast Probe PCR Kits
  • Multiplex probe detection: QuantiFast Multiplex PCR Kits

Ultrafast real-time two-step RT-PCR can be achieved when cDNA prepared with the FastLane Cell cDNA Kit is analyzed on the Rotor-Gene Q cycler using one of the following dedicated Rotor-Gene Kits:

  • SYBR® Green detection: Rotor-Gene SYBR® Green PCR Kit
  • Probe detection: Rotor-Gene Probe PCR Kit
  • Multiplex probe detection: Rotor-Gene Multiplex PCR Kit 

QuantiTect and QuantiFast Kits are compatible with various real-time cyclers, including instruments from Applied Biosystems.

アプリケーション

The FastLane Cell cDNA Kit is suitable for experiments which require a snapshot of individual transcript levels, such as:

  • Validation of siRNA-mediated gene knockdown
  • Evaluation of drug effects
  • Detection of gene regulation
Feature
Specifications
Applications Real-time, two-step RT-PCR, gene expression analysis directly from cells
Enzyme activity Reverse transcription
Mastermix No
Reaction type cDNA production, DNA digestion
Real-time or endpoint Real-time, two-step RT-PCR
Sample/target type Cultured cells
Single or multiplex Single
With/without hotstart Without hotstart

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キットハンドブック
3
For high-speed preparation of first-strand cDNA directly from cultured cells without RNA purification. For use in real-time, two-step RT-PCR
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RNA精製を行なわずに直接培養細胞から一本鎖cDNAを迅速に調製
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安全データシート
2
画像
Superior sensitivity in cancer research
がん研究において最高の感度を実現
HeLa細胞を用いたがん研究に重要な様々な遺伝子の2ステップリアルタイムRT-PCR解析において、Supplier AIVの類似キットの代わりにFastLane Kitを用いてcDNAを調製した時に、より低いCT値が得られた。
Successful analysis of CDC2 knockdown
CDC2ノックダウンの優れた解析結果

様々なCDC2 siRNAs(S1S2S3)あるいはnonsilencing siRNA(NS)をMCF-7細胞にトランスフェクトした。48時間後にFastLane Kitを用いてcDNAを調製した。GAPDH(内因性コントロール)に対するCDC2(細胞周期関連タンパク質)の相対的な発現を2ステップリアルタイムRT-PCRにより解析した。未処理細胞中(UT)の相対的なCDC2発現を100%とした。

Effective elimination of genomic DNA contamination
ゲノムDNAコンタミを効率的に除去
FastLane Cell cDNA Kitを用いて逆転写酵素を添加(+RT)あるいは未添加(-RT)でcDNAを調製した。β-actinの遺伝子発現アッセイを用いてcDNAを定量した。–RTの線は、ゲノムDNAが検出されなかったことを示唆している。
Highly reproducible cDNA preparation
再現性の高いcDNA調製
HeLa細胞を含む48ウェルプレートからFastLane Kitを用いてcDNAを調製した。Tubulin B(構造タンパク質)の発現を2ステップリアルタイムRT-PCRにより解析した。