A master mix type RT-PCR PreMix Kit capable of performing cDNA synthesis and PCR reaction at once using gene specific primers.
• One-step RT-PCR
• Ready to Use
– All components required for RT-PCR reaction are in a single tube with solution type
– Easy use by adding RNA, gene specific primers and DW only
• Sensitive cDNA synthesis and high specific PCR amplification
• Adjustable reaction volume according to requirement
• High specificity and accuracy by hot-start Taq DNA Polymerase
ONE-STEP RT-PCR PreMix Kit is a master mix type ONE-STEP RT-PCR PreMix Kit that can perform cDNA synthesis and PCR reaction all at once using gene specific primers. The general RT-PCR method is a two-step sequential reaction, cDNA synthesis using reverse transcriptase, oligo dT / random primers and PCR process with DNA polymerase. The method is suitable for gene expression studies, but it has drawbacks such as cross contamination in complex processes. Gene specific primer (GSP) is preferred for the confirmation of the existence of specific genes and quantitative / qualitative analysis of the expressed genes. It is important to minimize errors in the reaction by simplifying the experimental procedure. The ONE-STEP RT-PCR PreMix Kit was designed to synthesize first-strand cDNA from total RNA or mRNA template and do PCR in a tube. The product includes all components, reverse transcriptase and DNA polymerase, for each reaction with mixed solution type. The OptiScriptTM RT System of ONE-STEP RT-PCR PreMix Kit makes accurate cDNA and high specificity PCR from 1pg to 2mg of template RNA. The ONE-STEP RT-PCR PreMix Kit is optimized product to confirm the expression of low copy gene, and its stability is maintained for the first time under frozen condition for more than 1 year.
[ ONE-STEP RT-PCR PreMix Kit ]
Component in 20 ml reaction
• OptiScript™ RT System
• i-StarTaq™ DNA Polymerase
• RT-PCR Buffer(10x)
• dNTP Mixture
Observation of sensitivity and specificity through virus screening
After the decimal dilution of 108.0EID50 / 0.1ml of the uroepithelium, RNA was extracted with Viral Gene-spin™ Viral DNA / RNA Extraction Kit (Cat. No. 17151). Then first strand cDNA synthesis and PCR reaction were performed using ONE-STEP RT-PCR PreMix Kit. As can be seen from the result, it has high efficiency and specificity, so it can be used to detect various diseases.
Lane M, 100bp Ladder DNA Marker; lane 1, 10-2 희석; lane 2, 10-3 희석; lane 3, 10-4 희석; lane 4, 10-5 희석; lane 5, 10-6 희석; lane 6, 10-7 희석; lane 7, 10-8 희석; lane 8 : 10-9 희석
Observation of sensitivity and specificity
Total RNA was isolated from human cancer cell lines (SNU 5) using the easy-BLUE™ Total RNA Extraction Kit (Cat. No. 17061), and then synthesized each first strand of cDNA using AMV Reverse Transcriptase and Optiscript™ RT System. After the cDNA mixture was diluted to an appropriate concentration, the expression levels of β-Actin (400 bp) gene were compared. As can be seen from the above results, we found that Optiscript™ RT System is more sensitive than AMV RTase. Therefore, ONE-STEP RT-PCR PreMix Kit has high efficiency and specificity due to Optiscript™ RT System and i-StarTaq™ DNA Polymerase. In other words, OptiScript™ RT System allows sensitive reverse transcription reaction and i-StarTaq™ DNA Polymerase provides high specificity amplification as hot-start PCR enzyme.