QuantiTect SYBR^® Green RT-PCR Kit

For one-step qRT-PCR using SYBR Green I for gene expression analysis

High specificity in one-step RT-PCR
Reliable quantification of low-abundance transcripts
Accurate quantification over several logs of template
No need to optimize reaction and cycling conditions

The QuantiTect SYBR Green RT-PCR Kit provides highly specific quantification of RNA targets by real-time one-step RT-PCR using SYBR Green I detection. The combination of a hot start and a unique qRT-PCR buffer system ensures highly specific and sensitive real-time quantification of RNA targets. The dNTP mix includes dUTP, allowing optional treatment with UNG. The QuantiTect SYBR Green RT-PCR Kit is also supplied with an optimized RT mix for efficient and sensitive reverse transcription over a wide range of RNA template amounts. For convenience, the master mix can be stored at 2–8°C.

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製品名		Cat. no.	List price:
QuantiTect SYBR Green RT-PCR Kit (200) For 200 x 50 µl reactions: 3 x 1.7 ml 2x QuantiTect SYBR Green RT-PCR Master Mix, 100 µl QuantiTect RT Mix, 2 x 2 ml RNase-Free Water		204243	¥101,000
QuantiTect SYBR Green RT-PCR Kit (1000) For 1000 x 50 µl reactions: 25 ml 2x QuantiTect SYBR Green RT-PCR Master Mix, 0.5 ml QuantiTect RT Mix, 20 ml RNase-Free Water		204245	¥403,000

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QuantiTect SYBR^® Green RT-PCR 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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1ステップRT-PCR|特異性の高い増幅|1ステップおよび2ステップRT-PCRが類似した結果|特異的なプライマーアニーリング|

QuantiTect SYBR^® Green RT-PCR Kit は面倒で時間のかかる反応条件の至適化が不要。プライマーとテンプレートを即使用可能なRT-PCRマスターミックスに添加するだけで、ほとんどのリアルタイムサイクラーで反応を開始できる。|他社のDNAポリメラーゼと比較した結果、ユニークなバッファーと組み合わせたHotStarTaq DNA Polymeraseのみが497 bp フラグメントを特異的に増幅させた（1 µgのヒトゲノムDNAに添加した50 コピーのHIV-pol-遺伝子コンストラクトからのフラグメント）。M:マーカー。|HeLa細胞からのトータルRNA（10 ng～100 pg）あるいは匹敵する量のcDNAを、ヒトMAPK14用QuantiTect Primer Assay および表記のキットを用いて解析した。1ステップおよび2ステップRT-PCRで同等のC_T 値が得られた。|バランスの取れたKCl および(NH₄)₂SO₄ の組み合わせは、PCRテンプレートへのプライマーの特異的なアニーリングを促進する。K⁺ は二本鎖DNAのリン酸基に結合し、プライマーのアニーリングを安定化する。NH₄⁺ は、ミスマッチな塩基対間の水素結合を不安定化する。|

パフォーマンス

The QuantiTect SYBR Green RT-PCR Kit contains a unique blend of reverse transcriptases, which deliver efficient and sensitive cDNA synthesis over a wide range of amounts of RNA template and enables specific quantification over a wide linear range (see figure "Comparable results in one-step and two-step RT-PCR"). HotStarTaq DNA Polymerase further increases the specificity of the PCR reaction by providing the most stringent hot start compared with other polymerases (see figure "Highly specific amplification").

原理

QuantiTect SYBR Green RT-PCR Kits contain an optimized, ready-to-use master mix for highly specific and sensitive real-time quantification of RNA targets using SYBR Green I. The fluorescent dye SYBR Green I in the master mix enables the analysis of many different targets without having to synthesize target-specific labeled probes. A balanced combination of K⁺ and NH₄⁺ ions in the PCR buffer promotes specific primer annealing, enabling high RT-PCR specificity and sensitivity (see figure "Specific primer annealing"). In addition, an optimized mix of reverse transcriptases enables cDNA synthesis from a wide range of RNA template amounts, while HotStarTaq DNA Polymerase provides a stringent hot start, preventing the formation of nonspecific products.

QuantiTect SYBR Green RT-PCR master mix also contains dUTP, enabling pretreatment with uracil-N-glycosylase (UNG) prior to starting PCR, which ensures that any contaminating PCR products do not affect subsequent PCR reactions.

Components of 2x QuantiTect SYBR Green RT-PCR Kit
Component	Features	Benefits
HotStarTaq DNA Polymerase	15 min activation at 95ºC	Set-up of qPCR reactions at room temperature
QuantiTect SYBR Green RT-PCR Buffer	Balanced combination of NH₄⁺ and K⁺ ions	Specific primer annealing ensures reliable PCR results
dNTP mix	Includes dUTP, which partially replaces dTTP and enables optional UNG treatment of reactions	Eliminates contamination from carryover of PCR products by optional UNG treatment
SYBR Green I dye	Yields a strong fluorescent signal upon binding double-stranded DNA	Highly sensitive quantification
ROX dye	For normalization of fluorescent signals on Applied Biosystems and, optionally, Agilent instruments	Precise quantification on cyclers that require ROX dye. Does not interfere with reactions on other real-time cyclers
Omniscript and Sensiscript Reverse Transcriptases	Special blend of enzymes with high affinity for RNA	RNA can be transcribed, even through complex secondary structures

操作手順

The QuantiTect SYBR Green RT-PCR Kit overcomes the need for optimization of reaction conditions, which can be tedious and time-consuming. Simply add primers and template to the ready-to-use RT-PCR master mix, and start the reaction (see flowchart "One-step RT-PCR"). Follow the protocol in the handbook to get fast and reliable results on any real-time cycler. If required, reactions can be pretreated with uracil-N-glycosylase (UNG) to eliminate carryover of PCR products from previous reactions.

Highly specific results in gene expression analysis are guaranteed when QuantiTect SYBR Green RT-PCR Kits are used in combination with QuantiTect Primer Assays. These are genomewide, bioinformatically validated primer sets for detecting transcripts from human, mouse, rat, and many other species. QuantiTect Primer Assays can be easily ordered online at GeneGlobe.

アプリケーション

The QuantiTect SYBR Green RT-PCR Kit is for use in gene expression analysis of RNA targets. QuantiTect SYBR Green RT-PCR Kits are compatible with all available real-time cyclers, including instruments from Applied Biosystems, Bio-Rad, Cepheid, Eppendorf, Roche, and Agilent. For the Rotor-Gene Q and other Rotor-Gene cyclers, we recommend using the Rotor-Gene SYBR Green RT-PCR Kit, which has been specially developed for fast cycling on these instruments.

Feature	Specifications
Applications	Real-time quantification of RNA targets
Reaction type	Real-time and one-step RT-PCR
Real-time or endpoint	Real-time
Sample/target type	RNA
Single or multiplex	Single
SYBR Green I or sequence-specific probes	SYBR Green I
Thermal cycler	All real-time cyclers (e.g. LC, RG, ABI)
With or without ROX	With ROX

パンフレット

Sort options アルファベット順
	Flexible RNAi Technologies You Can Rely On - (EN)	詳細を表示
	(EN) - Analyzing Gene Expression and Regulation	詳細を表示
	(JA) - 信頼できる柔軟性の高いRNAi テクノロジー	詳細を表示
	(JA) - 遺伝子発現および発現制御の解析	詳細を表示
	Critical Factors for Successful Real-Time PCR - (EN)	詳細を表示

FAQ

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	Do you recommend 1-step or 2-step real-time RT-PCR for gene expression analysis? FAQ ID -1056	表示
	Why does my PCR product show up later when comparing the QuantiTect SYBR Green PCR Kits with Roche kits using the same annealing temperature? FAQ ID -1083	表示
	Why do melting temperatures differ between PCR fragments amplified with QIAGEN's QuantiTect SYBR Green PCR Kits and Roche Kits? FAQ ID -1084	表示
	What should I use as a standard for absolute quantification in real-time PCR? FAQ ID -1085	表示
	How much template can I use in the reaction and what is the maximum volume of template that can be used in the QuantiTect Kits? FAQ ID -1086	表示
	What is a QuantiTect Primer Assay? FAQ ID -1141	表示
	Do QuantiTect Primer Assays contain SYBR Green dye? FAQ ID -1143	表示
	Can I increase the annealing temperature recommended for QuantiTect Primer Assays used with the QuantiTect SYBR Green PCR Kits? FAQ ID -1146	表示
	How much time will be saved when switching from standard cycling to fast cycling with QuantiFast Kits? FAQ ID -1438	表示
	How do QuantiFast PCR Kits compare to QuantiTect PCR Kits for quantitative real-time PCR? FAQ ID -1441	表示
	Why is the storage time for QuantiFast PCR Kits shorter than that for QuantiTect PCR Kits? FAQ ID -1446	表示
	Why is the reaction volume for QuantiFast PCR Kits lower than that for QuantiTect PCR Kits? FAQ ID -1447	表示
	Why is the RT step with the QuantiFast RT Kits much shorter compared to QuantiTect RT Kits? FAQ ID -1451	表示
	Have QuantiTect Primer Assays been tested with QuantiFast SYBR Green PCR Kits on the Mastercycler ep realplex? FAQ ID -1714	表示
	QuantiTect Primer Assays are bioinformatically validated, genomewide primer sets. What does “bioinformatically validated” mean? FAQ ID -1982	表示
	Why is an ice-incubation step included during reaction set-up when following the QuantiTect RT-PCR but not the QuantiTect PCR protocol. FAQ ID -283	表示
	Can Uracil-N-Glycosylase (UNG) be purchased separately, independent of QuantiTect PCR Kits? FAQ ID -2134	表示
	Can Uracil-N-Glycosylase (UNG) be used with a one-step RT-PCR kit, such as the QuantiTect Probe RT-PCR Kit? FAQ ID -2135	表示
	Do you have any information or guidelines regarding the choice of reference genes for real-time PCR? FAQ ID -2371	表示
	How do I create a workspace that is free of DNA contamination, prior to carrying out a qPCR experiment? FAQ ID -2654	表示
	How important is the RNA purification process, for obtaining reliable qRT-PCR results? FAQ ID -2655	表示
	What is the threshold cycle or Ct value? FAQ ID -2682	表示
	Does SYBR Green dye present in QuantiTect kits interfere with ethidium bromide staining? FAQ ID -312	表示
	Do the QuantiTect SYBR Green Kits contain a stabilization reagent that inhibits the sensitivity of real-time PCR? FAQ ID -328	表示
	My QuantiTect Master Mix did not freeze at degrees. Is it still ok to use? -20	表示
	Why do replicates in real-time PCR have different plateau heights? FAQ ID -539	表示
	What reaction volume should I use with QuantiTect Kits? FAQ ID -551	表示
	How long should the amplicon be if I am using QuantiTect SYBR Green detection? FAQ ID -553	表示
	Is it possible to perform a UNG treatment when using QuantiTect kits? FAQ ID -564	表示
	Do you have information on the use of recombinant DNA and RNA as absolute standards for realtime RT-PCR? FAQ ID -729	表示
	Why should DNA or cDNA targets be less than 250 bp long for real-time PCR? FAQ ID-751	表示
	What annealing temperature should be used with the QuantiTect Primer Assays? FAQ ID -849	表示
	What is the concentration of the primers in a reaction using the QuantiTect Primer Assays? FAQ ID -850	表示

キットハンドブック

Sort options アルファベット順
	(EN) - QuantiTect Primer Assay Handbook For genomewide, ready-to-use real-time RT-PCR assays using SYBR Green detection	詳細を表示
	(EN) - QuantiTect SYBR Green RT-PCR Handbook For quantitative, real-time one-step RT-PCR using SYBR Green I	詳細を表示
	(JA) - QuantiTect SYBR Green RT-PCRプロトコールとトラブルシューティング SYBR Green I を用いたリアルタイム定量1ステップRT-PCR 用	詳細を表示

サイエンティフィック・ポスター

Sort options アルファベット順
	Quantitative RT-PCR analysis with genomewide, validated primer sets - (EN)	詳細を表示

テクニカルインフォメーション

Sort options アルファベット順
	(JA) - 簡便で正確なリアルタイムPCR実現のために	詳細を表示
	(JA) - SYBR Green マスターミックスの重要性	詳細を表示

MSDS

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	Safety Data Sheets Download Safety Data Sheets for QIAGEN product components.	表示

安全データシート

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	MSDS QuantiTect SYBR Green RT-PCR Kit (200)
	MSDS QuantiTect SYBR Green RT-PCR Kit (1000)
	CofA

画像

1ステップRT-PCR

QuantiTect SYBR^® Green RT-PCR Kit は面倒で時間のかかる反応条件の至適化が不要。プライマーとテンプレートを即使用可能なRT-PCRマスターミックスに添加するだけで、ほとんどのリアルタイムサイクラーで反応を開始できる。

特異性の高い増幅

他社のDNAポリメラーゼと比較した結果、ユニークなバッファーと組み合わせたHotStarTaq DNA Polymeraseのみが497 bp フラグメントを特異的に増幅させた（1 µgのヒトゲノムDNAに添加した50 コピーのHIV-pol-遺伝子コンストラクトからのフラグメント）。M:マーカー。

Comparable results in one-sep and two-step RT-PCR on the LightCycler 2.0.

1ステップおよび2ステップRT-PCRが類似した結果

HeLa細胞からのトータルRNA（10 ng～100 pg）あるいは匹敵する量のcDNAを、ヒトMAPK14用QuantiTect Primer Assay および表記のキットを用いて解析した。1ステップおよび2ステップRT-PCRで同等のC_T 値が得られた。

Specific primer annealing using a balanced combination of K+ and NH4+ ions

特異的なプライマーアニーリング

バランスの取れたKCl および(NH₄)₂SO₄ の組み合わせは、PCRテンプレートへのプライマーの特異的なアニーリングを促進する。K⁺ は二本鎖DNAのリン酸基に結合し、プライマーのアニーリングを安定化する。NH₄⁺ は、ミスマッチな塩基対間の水素結合を不安定化する。

QuantiTect SYBR® Green RT-PCR Kit

QuantiTect SYBR^® Green RT-PCR Kit