● Purity: Reverse transcription and PCR reactions are completed in one step to avoid cross contamination. 

● High efficiency: Unique King reverse transcriptase with RT efficiency over 95%. 

● Sensitive: As low as 1 ng templates can be accurately identified, especially for templates with low abundance. 

● Specificity: The antibody-modified Taq polymerase further improves the amplification efficiency and specificity.

FastKing One Step RT-PCR Kit allows both reverse transcription and gene amplification to take place in a single tube, which avoids cross contamination between samples and improves the sensitivity of detection. The 25× RT-PCR Enzyme Mix contains King reverse transcriptase, which is a high efficient reverse transcriptase expressed by engineering bacteria; a further-modified hot start Taq DNA polymerase, which provides high efficiency and accuracy for the amplify reaction; and RNase inhibitor. With a special modified hydrophobic motif, King RTase gets a significant affinity for RNA and facilitates transcription through of RNA templates, especially for templates with high GC content or complex secondary structures. The 2× FastKing One Step RT-PCR MasterMix contains appropriate ion concentration, dNTPs and PCR enhancer. It could stabilize both RTase and polymerases and keep their efficiency within the whole reaction process.

● RNA template could be total RNA or mRNA. TRNzol or RNAprep pure kits can be used to purify high-quality total RNA.

● RNase contaminations should be avoided in one-step RT-PCR. Some measures can be taken as below:

    • Wear a disposable gloves and respirator to avoid the RNase contaminations from skin and saliva.

    • Operate the RNA related experiments in an RNase-free environment using RNase-free apparatus and consumable items.

    • Consumable items related with RT-PCR should be incubated in 0.1% DEPC solution at 37°C for 12 hours and sterilized for 30 min before use.

● 25× RT-PCR Enzyme Mix should be centrifuged briefly before use. Pipet slowly and store at -30~-15°C immediately after use.

● Completely mix the 2× FastKing One Step RT-PCR MasterMix and centrifuge the reagent to the botiom of the tube before use.

● Only use gene-specific primers and use specific primers according to specific requirements of experiments.

● RNA templates

● Gene-specific PCR primers

● Thoroughly thaw the template RNA, gene-specific PCR primers, 2× FastKing One Step RT-PCR MasterMix and RNase-Free ddH₂O, centrifuge briefly and place them on ice.

● Prepare a reaction solution according to the following table on ice:

● Set up thermal cycler conditions according the following table.

● Analyze the PCR products using agarose gel electrophoresis.