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Some experience about siRNA in RT-PCR
1. Basic knowledge (if you want to see the experimental part, please transfer directly to the second part) As a derivative reaction of conventional PCR, Real time PCR mainly monitors the change of the amount of amplification product in each cycle of the PCR amplification reaction in real time thr...Read more -
RT-PCR experimental reaction system optimization method detailed summary
一、Increase the sensitivity of the reaction system： 1. Isolate high quality RNA： Successful cDNA synthesis comes from high-quality RNA. High-quality RNA should at least be full-length and free of reverse transcriptase inhibitors such as EDTA or SDS. The quality of RNA determines the maximum a...Read more -
Complete PCR primer design and PCR details
Primer design basis (99% problems can be solved) 1. Primer length: The textbook requires 15-30bp, usually about 20bp. The actual condition is better to be 18-24bp to ensure the specificity, but the longer the better, too long primer will also reduce the specificity, and reduce the yield. 2. Prim...Read more -
Precautions for qRT-PCR compiled by senior Doctor painstakingly
Everyone is talking about the principle of qRT-PCR experiment, primer design, result interpretation, etc., but I think I should share with you the experimental operation of qRT-PCR. It’s small, but it’s about results. Before doing qRT-PCR, we need to have a clear understanding of our ...Read more -
Precautions for RT-qPCR
RT-qPCR experiment includes RNA extraction and quality assessment, reverse transcription and qPCR three steps, each step has a lot of precautions, we will introduce in detail below. Ⅰ. RNA quality assessment In RT-qPCR experiment, after the completion of RNA extraction, ...Read more -
A thorough introduction to real time PCR
1. Initial understanding At this stage, we need to understand some concepts and terminology, so as to avoid making mistakes in front of our seniors, such as: Q: What is the difference between RT-PCR, qPCR, Real-time PCR, and real-time RT-PCR? Answer: RT-PCR is reverse transcription PCR (reverse t...Read more -
PCR, multiple PCR, In situ PCR, Reverse PCR, RT-PCR, qPCR（1）– PCR
PCR, multiple PCR, In situ PCR, Reverse PCR, RT-PCR, qPCR（1）– PCR We will sort out the concepts, steps and details of various PCR Ⅰ. PCR Polymerase Chain Reaction, referred to as PCR, is a molecular biological technology that is used to enlarge specific DNA fragments. It can be regarded a...Read more -
qRT-PCR experimental principle
RT-qPCR is developed from ordinary PCR technology. It adds fluorescent chemicals (fluorescent dyes or fluorescent probes) to the traditional PCR reaction system, and detects the PCR annealing and extension process in real time according to their different luminescent mechanisms. Fluorescent signa...Read more -
RT-PCR experimental reaction system optimization method detailed summary
Ⅰ. Increase the sensitivity of the reaction system: 1. Separate high -quality RNA: Successful cDNA synthesis comes from high -quality RNA. High -quality RNA should ensure at least a total longer and does not contain inhibitors that do not contain recording enzymes, such as EDTA or SDS. The quali...Read more -
How To Sure Reasonable Range Of Ct Values For qPCR Experiments
Ct value is the most important result presentation form of fluorescent quantitative PCR. It is used to calculate gene expression differences or gene copy number. So what is the Ct value of fluorescence quantification considered reasonable? How to ensure the effective range of Ct value? What is Ct...Read more -
Pay attention to the following details to double the success rate of your RT-qPCR experiment!
RT-qPCR is the basic experiment of molecular biology, and everyone must be familiar with it. It mainly includes three steps: RNA extraction, reverse transcription into cDNA, and real-time fluorescent quantitative PCR. It doesn’t help, what is going on? It is likely that there is a problem w...Read more -
RNA extraction, the 260/230 is particularly low, and the ct value of pcr run is very high. How to solve it?
The low ratio of A260/A230 is usually caused by impurities with the maximum absorption wavelength at 230nm. Let’s see what these impurities include: Common pollutants Absorption wavelength Ratio effect Protein ~230nm and 280nm Simultaneous reduction of A 260 /A 280 and A 260 /A ...Read more
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