SYBR Green qPCR Mix (Rox+)

The fluorescent substances used for real-time fluorescence quantitative PCR can be divided into two types: fluorescent probes and fluorescent dyes. The principle is briefly described as follows:

1. TaqMan fluorescent probe: A specific fluorescent probe is added at the same time that a pair of primers is added during the PCR amplification. The probe is an oligonucleotide, and a reporter fluorescent group and a quencher are labeled on both ends respectively. Fluorescent group. When the probe is intact, the fluorescent signal emitted by the reporter group is absorbed by the quencher group; when PCR is amplified, the Taq enzyme's 5′-3′ exonuclease activity degrades the probe, making the reporter fluorescent group and quenching. The fluorescein group is separated, so that the fluorescence monitoring system can receive the fluorescence signal, that is, one fluorescent molecule is formed for each DNA strand to be amplified, and the accumulation of the fluorescence signal is completely synchronized with the formation of the PCR product. The new TaqMan-MGB probe enables this technology to perform both quantitative gene analysis and gene mutation (SNP) analysis, and is expected to be the preferred technology platform for genetic diagnosis and individualized drug analysis.

2. SYBR fluorescent dyes: In the PCR reaction system, excess SYBR fluorescent dye is added. SYBR fluorescent dyes non-specifically incorporate DNA double strands, emit fluorescent signals, and SYBR dye molecules that do not incorporate into the chain do not emit any fluorescence. The signal is thus ensured that the increase of the fluorescence signal is completely synchronized with the increase of the PCR product. SYBR only binds to double-stranded DNA, so it is possible to determine whether the PCR reaction is specific by the dissolution profile.

3. Molecular Beacon: A stem-loop double-labeled oligonucleotide probe that forms a hairpin structure of approximately 8 bases at its 5 and 3 ends itself, with complementary nucleic acid sequences at both ends, resulting in a fluorophore group Quencher groups are close together and do not produce fluorescence. After the PCR product is generated, during the annealing process, the middle portion of the molecular beacon is paired with the specific DNA sequence, and the fluorescent gene is separated from the quencher gene to generate fluorescence.

Method Advantages Disadvantages Scope

SYBR Green I Wide range of applications, sensitive, convenient, cheap primers, prone to non-specific bands in the quantitative analysis of multiple target genes, gene expression research

Taq Man has high specificity, good reproducibility and high price, it is only suitable for specific target pathogen detection, disease resistance gene research, drug efficacy assessment, genetic disease diagnosis

High Specificity of Molecular Beacons (HRM), High Distinction, High Requirements for Instruments and Reagents, and Study of Single-base Mutations such as SNPs

This article will introduce the newly developed SYBR Green qPCR Mix from Guangzhou Dongsheng Biological. The DNA polymerase in this product is an improved Taq DNA polymerase, combined with the newly developed Buffer system, which can effectively inhibit non-specific PCR amplification. It greatly increases the sensitivity of PCR amplification. At the same time, Rox internal reference dye is added to the reaction system, which can correct fluorescence signal errors between different wells and is suitable for high-sensitivity Real Time PCR amplification reactions. This product is suitable for rapid real-time PCR amplification reaction, can get a good standard curve in a wide range of quantitative areas, accurate quantification of target genes, detection, good repeatability, high credibility.

Rox+ dye is a fluorescent dye that does not participate in the PCR reaction. The maximum absorption wavelength is about 576nm, and the maximum emission wavelength is about 601nm. Its main function is to correct the sample amount error and the fluorescence signal of each well passes through the filter, and then passes through Focus on the difference in the optical path of the CCD.

Features

? High Specificity: Hot Start Function and Optimized Buffers Prevent Nonspecific Amplification and Primer Dimer Formation

? Sensitivity: detectable low copy template

? Wide linear range: accurate quantification of 9 orders of magnitude

? Versatility: Almost compatible with all real-time PCR instruments

? Reproducibility, easy to use: ready-to-use 2×Mix, reducing sample loading errors and reaction system configuration time

Product Usage

? Real-time PCR (using SYBR® GreenI)

? Real-time RT-PCR (using SYBR® GreenI)