We are also doing PCR experiments, why are the results always so much worse than others?

We are also doing PCR experiments, why are the results always so much worse than others?

In the laboratory, look at other people’s PCR, amplification efficiency is high, electrophoresis strip perfect, and look at their own, hard to do for a long time, feel the reaction is very good, the result of a false positive test…… What’s the problem?

I believe that many of our partners have also encountered various problems. When we run into a wall in PCR experiments, we will subconsciously analyze the factors within the reaction system, but we often ignore the important influence of consumables in experiments.

Your PCR result is wrong, the PCR consumables may be the essential reason. On the one hand, micro contamination of consumables or the introduction of inhibitors will cause experimental pollution; On the other hand, improper selection of consumables will also make the experimental results suffer.

The problem comes again. There are many types of PCR consumables, so how to choose the right ones to make the experiment more smooth? What are the problems to be paid attention to? Don’t worry, today I combined with many years of experience, sorting out the PCR consumables selection common problems, I hope to help you.

 

Q1: Why are PCR consumables usually made of PP material?

Huida: Because PCR supplies are usually in direct contact with reagents or samples, polypropylene (PP) materials are biologically inert, do not adhere to biomoleculars, and have good chemical tolerance and temperature tolerance (can be autoclapped at 121 ° C, can also withstand temperature changes during the thermal cycle).

 

Q2: How should I choose between different sizes of PCR tubes/plates?

HUIDA: Purpose of selection: select the right products according to the specific experimental requirements. Most of the size of the PCR tube can meet the requirements of PCR reaction. On this basis, it is recommended to choose the low volume tube first. Because the low volume reaction tube/plate has a small overhead space, the heat conductivity is improved and evaporation is reduced. At the same time, it is necessary to avoid adding too much or too little samples. Too much can lead to decreased thermal conductivity, spillage and cross-contamination, while too little may result in sample evaporation loss.

Specifications and volumes of common reaction tubes:

Single/combined tube: 0.5ml, 0.2ml, 0.15ml

96-well plate: 0.2ml, 0.15ml

384-well plate: 0.04ml

 

Q3: When the sample quantity is small, I will choose single pipe or combined pipe, but why do some have flat cover and some have convex cover?

Huida: When the sample quantity is small, the single tube or the combined tube will be preferred. However, when the reaction volume is large, the single tube is dominant, and the volume can reach 0.5 mL. And flat cover and convex cover each has the advantage, specific can refer to the following information.

Single tube: 0.2ml and 0.5ml; Can choose sample quantity flexibly.

Connecting tube: 0.2ml or 0.15ml is optional; 8 or 12 tubes are common.

Flat cover: can provide accurate fluorescence signal transmission for qPCR; Easy to write marks.

Convex cover: contact with the hot cover of the PCR instrument to reduce the deformation of the reaction tube caused by pressure; However, it will affect the fluorescence signal transmission and cannot be applied to the qPCR experiment.

 

Q4: Why do some PCR plates have skirts and some don’t?

Huida: In fact, the skirt of the PCR plate is designed to better adapt to automation applications, providing stable support and mechanical endurance for the instrument, as well as higher stability during the pipetting process.

PCR plates are generally divided into non-hemline, half-hemline and full-hemline.

Without Plate: Suitable for most PCR machines or qPCR machines, but not for automation applications. The stability is not high in the process of pipetting, so it needs to be used with plate support.

Half skirt edge plate: can be adapted to label or bar code application, and automatic application, and has good pipetting stability.

Full skirt edge plate: very suitable for automated experimental applications, can also be adapted to the label and application of bar code. It has good mechanical strength, can be used in bulge module PCR instrument, and has high stability in the process of pipetting.

 

Q5 :For PCR plates, why are the corners and labels not quite the same?

Huida: It depends on the effect of the corner cutting and the marking.

Corner cutting: The choice of corner cutting position of PCR plate depends on the requirements of the suitable instrument for easy positioning.

Identification: Alphanumeric markers on PCR plates help identify individual Wells and sample locations. Generally for the embossed color digital logo or engraved logo. For some automated applications, printed and marked reaction plates are better sealed.

 

Q6: Why are some PCR plates convex while others are flat? Which is better?

Huida: What fits is best. There are two kinds of common PCR plates: flat pore margin and high pore margin.

Flat hole margin reaction plate, suitable for most PCR apparatus.

Reaction plates with rising orifice edges facilitate membrane sealing and reduce the risk of cross-contamination between samples.

 

Huida has many years of experience in the production and processing of PCR products and industry high standards. The product line is rich and can meet the vast majority of PCR experimental applications. Come and choose the right products for your experimental applications.

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Post time: Jun-03-2021