Description 

The ExcelRT™ One-Step RT-qPCR kit (TaqMan, no ROX) is designed for reverse transcription and quantitative real-time analysis of a specific target RNA by one-step reaction. The ExcelRT™ One-Step RT-qPCR kit (TaqMan, no ROX), consisting of One-Step RT Enzyme Mix and 2X One-Step Master Mix, is a convenient kit designed for highly efficient cDNA synthesis and highly specific real-time PCR in a single tube. The One-Step RT Enzyme Mix contains a thermostable ExcelRT™ Reverse Transcriptase and a RNAok™ RNase inhibitor. Consequently, One-Step RT Enzyme Mix can reverse transcribe RNA to cDNA at a wide temperature range from 42 to 60°C and active against RNase A, RNase B and RNase C. By containing specialized hot-start Taq DNA polymerase, which greatly reduce primer-dimer formation and can be activated within 2 minutes, the 2X One-Step Master Mix features high specificity and is suitable for fast cycle program.

Features

• Reverse transcription at wide temperature range (42-60°C)
• High specificity 
• Suitable for fast cycle program
• With no ROX reference dye

Storage

Aliquot to avoid multiple freeze-thaw cycles (stable within 30 freeze-thaw cycles)

Protect from light

-20°C for 12 months

The ExcelRT™ One-Step RT-qPCR kit (TaqMan, no ROX) is designed for reverse transcription and quantitative real-time analysis of a specific target RNA by one-step reaction. The ExcelRT™ One-Step RT-qPCR kit (TaqMan, no ROX), consisting of One-Step RT Enzyme Mix and 2X One-Step Master Mix, is a convenient kit designed for highly efficient cDNA synthesis and highly specific real-time PCR in a single tube. The One-Step RT Enzyme Mix contains a thermostable ExcelRT™ Reverse Transcriptase and a RNAok™ RNase inhibitor. Consequently, One-Step RT Enzyme Mix can reverse transcribe RNA to cDNA at a wide temperature range from 42 to 60°C and active against RNase A, RNase B and RNase C. By containing specialized hot-start Taq DNA polymerase, which greatly reduce primer-dimer formation and can be activated within 2 minutes, the 2X One-Step Master Mix features high specificity and is suitable for fast cycle program.

Introduction

The Kit integrates phenol/guanidine-based lysis and silica membrane purification of total RNA. MagZol Reagent, included in the kits, is a monophasic solution of phenol and guanidine thiocyanate, designed to facilitate lysis of samples and inhibit RNases. The high lysis efficiency of the reagent and the subsequent removal of contaminants by organic phase extraction enable extracting up to 200μg total purified RNA from less than 100mg animal, plant, fungal samples,bacteria, cells and other samples.

Details

Specifications

Features	Specifications
Main Functions	Isolation total RNA (include miRNA) from 100mg lipid tissue, tissue, cell, plant, body fluids using columns and MagZol reagent
Applications	RT-PCR, qRT-PCR, Northern hybridization, second generation sequencing
Purification method	Mini spin column
Purification technology	Silica technology, acid phenol / guanidine extraction technology (MagZol pretreatment technology)
Process method	Manual (centrifugation or vacuum)
Sample type	Animal tissue, muscle fiber containing tissue,adipose tissue, cultured cells, lymphocytes, simple plants and other biological samples
Sample amount	Animal tissue sample: 1-60mg (spleen≤ 10mg)Cultured cells: 5 x 106 Plant leaves: 50-100mg
Yield	2-200μg
Elution volume	≥50μl
Time per run	≤30 minutes（1-24 samples）
Liquid carrying volume per column	800µl
Binding yield of column	100µg

Principle

Tissue samples (10-100mg) are homogenized in MagZol Reagent. After addition of chloroform, the homogenate is separated into aqueous and organic phases by centrifugation. The upper, aqueous phase is extracted, and ethanol is added to provide appropriate binding conditions. The sample is then applied to the spin column, where the total RNA (up to 100µg) binds to the membrane and phenol and other contaminants are efficiently washed away. High-quality RNA is then eluted in 30-100µl of RNase-free water.

Advantages

• Efficient DNA removal – one step RNA extraction can effectively remove genomic DNA
• High quality – one step RNA extraction reagent combined with silica gel column can obtain the highest concentration
• Fast – several samples can be extracted in 30 minutes
• High applicability – samples including animals, plants, bacteria, cells, etc.
• High output – enable to process large samples and the yield can be up to 200μg

Kit Contents

Contents	R413002	D413003
Purification Times	50 Preps	250 Preps
HiPure RNA Mini Columns	50	250
2ml Collection Tubes	50	250
MagZol Reagent	60 ml	270 ml
Buffer RW1	50 ml	200 ml
Buffer RW2*	20 ml	50 ml
RNase Free Water	10 ml	30 ml

Storage and Stability

MagZol Reagent should be stored at 2-8°C upon arrival. However, short-term storage (up to 12 weeks) at room temperature (15-25°C) does not affect its performance. The remaining kit components can be stored at room temperature (15-25°C) and are stable for at least 18 months under these conditions.

Experiment Data

The Kit integrates phenol/guanidine-based lysis and silica membrane purification of total RNA. MagZol Reagent, included in the kits, is a monophasic solution of phenol and guanidine thiocyanate, designed to facilitate lysis of samples and inhibit RNases. The high lysis efficiency of the reagent and the subsequent removal of contaminants by organic phase extraction enable extracting up to 200μg total purified RNA from less than 100mg animal, plant, fungal samples,bacteria, cells and other samples.