Qiagen, 205313, QuantiTect Rev. Transcription Kit (200)

Name: QuantiTect Rev. Transcription Kit (200)
Brand: Qiagen
SKU: 205313

For 200 x 20 µl reactions: 4 x 100 µl 7x gDNA Wipeout Buffer, 4 x 50 µl Quantiscript Reverse Transcriptase, 4 x 200 µl 5x Quantiscript RT Buffer, 4 x 50 µl RT Primer Mix, 4 x 1.9 ml RNase-Free Water

Catalog Number: 205313

Pricing

Send an RFQ.

View RFQ List

Category

Reverse Transcriptases

TagsDiscovery & Translational Research PCR/qPCR/dPCR Real-Time PCR Enzymes & Kits Reverse Transcription & cDNA Synthesis for qPCR

Description

The unique QuantiTect Reverse Transcription Kit provides a fast and convenient procedure for cDNA synthesis with integrated genomic DNA removal. Genomic DNA contamination in RNA samples is effectively eliminated by gDNA Wipeout Buffer. All components that are required for fast and efficient reverse transcription are provided with the QuantiTect Reverse Transcription Kit, including Quantiscript Reverse Transcriptase, Quantiscript RT Buffer, and a unique RT Primer Mix. The synthesized cDNA is optimized for use in real-time PCR, allowing reliable quantification of targets from all regions of mRNA transcripts.
Using the QuantiTect Reverse Transcription Kit, contaminating genomic DNA in RNA samples is effectively and rapidly removed with the unique gDNA Wipeout Buffer (see figure Effective genomic DNA removal for accurate real-time RT-PCR). Elimination of genomic DNA is crucial for accurate gene expression results, and design of RNA-specific primers or probes is not always possible. With gDNA Wipeout Buffer, time is saved and costs are reduced, since a separate DNase digestion is unnecessary, either during or after purification of RNA samples. The high RNA affinity of Quantiscript Reverse Transcriptase, in combination with Quantiscript RT Buffer, enables high yields of cDNA from any RNA template (see table “Higher cDNA yields for less abundant transcripts”). Even difficult templates, such as those with high GC-content or complex secondary structure, are successfully reverse transcribed. Higher cDNA yields for less abundant transcripts C T values for IL12A (low expression) C T values for IL1RN (higher expression) Input RNA (ng) QIAGEN Supplier A II QIAGEN Supplier A II 1000 30.9 32.0 23.1 24.9 100 34.2 35.4 26.3 26.6 10 37.8 46.8 29.7 30.3 1 N.D. N.D. 32.4 34.5 Real-time, two-step RT-PCR analysis of IL12A and IL1RN using different amounts of input RNA. Total RNA was reverse transcribed using the QuantiTect Reverse Transcription Kit or a kit from Supplier A II. The cDNA synthesized was analyzed on the ABI PRISM 7900 using the QuantiTect Probe PCR Kit and QuantiTect Gene Expression Assays for IL12A or IL1RN. C T values were lower with the QIAGEN kit, especially for the medium-abundance IL12A gene (bold), indicating higher cDNA yields. N.D.: Not detected. The RT Primer Mix contains a specially optimized mix of oligo-dT and random primers that enable cDNA synthesis from all regions of RNA transcripts, even from 5' regions (see figure Sensitive detection of a target at the 5' region of a 12.5 kb transcript). In contrast to kits from other suppliers, the QuantiTect Reverse Transcription Kit provides high yields of cDNA template for real-time PCR analysis regardless of where the amplified target region is located on the transcript, and provides greater sensitivity in the detection of low-abundance genes (see figure "Higher sensitivity in real-time, two-step RT-PCR"). The QuantiTect Reverse Transcription Kit also enables greater reproducibility in real-time RT-PCR.
QuantiTect Reverse Transcriptase is a novel blend of Omniscript and Sensiscript Reverse Transcriptases, which has a high affinity for RNA and is capable of cDNA synthesis from a wide range of RNA amounts (10 pg to 1 µg). In contrast to kits from other suppliers, the QuantiTect Reverse Transcription Kit provides high yields of cDNA template for real-time PCR analysis regardless of where the amplified target region is located on the transcript. Even difficult templates, such as those with high GC-content or complex secondary structure, are successfully reverse transcribed. QuantiTect RT Buffer has also been optimized to be compatible with real-time PCR buffer. To obtain accurate results in real-time RT-PCR gene expression assays, it is important that only cDNA is amplified and detected. Interference by genomic DNA can be avoided by designing primers or probes that span an exon/exon boundary. However, in cases where this is not possible (e.g., the cDNA is from a single-exon gene), it is essential that the starting RNA sample is free of genomic DNA. Using the QuantiTect Reverse Transcription Kit, contaminating genomic DNA in RNA samples is effectively and rapidly removed with unique gDNA Wipeout Buffer. Time is saved and costs are reduced, since a separate DNase digestion not required, either during or after purification of RNA samples. Also, design of RNA-specific primers or probes is unnecessary. Components of the QuantiTect Reverse Transcriptase Kit Component Benefits gDNA Wipeout Buffer Detection of RNA only in real-time RT-PCR Quantiscript Reverse Transcriptase Use of a wide range of RNA amounts (10 pg to 1 µg RNA) High sensitivity Quantiscript RT Buffer Read-through of difficult templates RT Primer Mix cDNA synthesis from all regions of transcripts, even from 5' regions
Genomic DNA removal and cDNA synthesis take only 20 minutes with the QuantiTect Reverse Transcription Kit (see flowchart "Fast and convenient cDNA synthesis"). The procedure is fast and convenient, since both reactions are run using the same incubation temperature and are set up using master mixes. The QuantiTect Reverse Transcription Kit includes everything you need for fast cDNA synthesis. Purified RNA is briefly incubated in gDNA Wipeout Buffer to effectively remove contaminating genomic DNA. In contrast to other methods, the RNA sample is then used directly in reverse transcription, using a master mix prepared from Quantiscript Reverse Transcriptase, Quantiscript RT Buffer, and RT Primer Mix. With Quantiscript Reverse Transcriptase, RNA can be transcribed at low temperatures, even through complex 2° structure, ensuring that the RNA will stay intact — the entire reaction takes place at 42°C and is then inactivated at 95°C. Additional steps for RNA denaturation, primer annealing, and RNase H digestion are not necessary.

Specifications

General

Applications	Quantification of (even low-abundance) transcripts
Sample/target type	RNA template
Enzyme activity	Reverse transcription
Real-time or endpoint	Real time
Reaction type	Two-step, cDNA production, genomic DNA digestion
Single or multiplex	Single
Mastermix	No

FAQs

Question: How many reverse-transcription reactions are included?

Answer:This configuration provides reagents for 200 standard reactions.

Question: How is genomic DNA addressed before cDNA synthesis?

Answer:The workflow uses gDNA Wipeout Buffer before the reverse-transcription step.

Question: What RNA input range is supported for a standard reaction?

Answer:A standard 20 µL reaction can use approximately 10 pg to 1 µg total RNA.

Application Note

QuantiTect® Reverse Transcription Kit is used to convert RNA into cDNA for quantitative real-time PCR and other downstream amplification workflows. The kit combines genomic-DNA removal and reverse transcription in a two-step protocol. gDNA Wipeout Buffer first removes contaminating genomic DNA, after which Quantiscript Reverse Transcriptase, Quantiscript RT Buffer and RT Primer Mix generate first-strand cDNA. The reverse-transcription enzyme mixture is based on Omniscript and Sensiscript reverse transcriptases and includes RNase inhibitor. A standard reaction volume is 20 µL and can use approximately 10 pg to 1 µg total RNA. This configuration supplies reagents for 200 standard reactions, including four sets of the main components. The workflow is useful for gene-expression studies where genomic-DNA carryover could otherwise contribute to qPCR background.

Ordering

Notice

Unless otherwise stated, the minimum order amount is USD 1,000.
For orders below USD 1,000, an additional handling fee may apply, or we may arrange local delivery / local handover where available.
All quotations are subject to final confirmation, including product availability, lead time, batch information, shipping cost, and applicable export or handling requirements.
For quotation requests, please send an RFQ from website or contact:

[email protected]

Please send inquiries directly to us in the “To” field. Emails sent via BCC, mass mailing, or with unrelated recipients may be automatically filtered and will not be processed.