Choose the right tools
The preparation of the PCR is an essential step to achieve good results. In order to increase the speed of preparation, the master mix is mixed in a larger vessel and a multipette is used instead of a pipette to distribute the master mix into the reaction vessels. The entire liquid is absorbed and the right amount is dispensed into the tubes in increments. This works particularly well when the throughput is also increased and tube strips, 96-well plates, or even 384-well plates are used instead of individual vessels.
Tip: When using 96- and 384-well plates, it is worth considering purchasing a small pipetting station
. The reproducibility when preparing the master mix is increased, it is far less tiring for you and you have time to do something else while the robot is pipetting.
Another way to speed up the PCR is to use " Fast PCR tubestrips
" - filling takes the same time, but due to the extremely thin walls, the temperature in the sample is reached faster and the PCR itself takes less time.
The first thing that comes to mind to speed up a PCR is a faster cycler. Silver blocks are faster than aluminum blocks...but not always. It is important to compare how the devices are heated and this is largely due to the quality and quantity of the Peltier elements installed. For example, the Mastercycler® X50a
with an aluminum block is faster than some other cyclers with a silver block.
Are the optimization of PCR protocols and the frequent use of new primers important to you, a gradient, better still a 2D gradient in the cycler must be possible. A 2D gradient
enables the simultaneous optimization of annealing and denaturation temperatures in order not only to increase primer binding, but also to increase yield. Here, 96 or 384 different annealing and denaturing temperatures are tested in combination within one PCR run.
If there are only a few samples, it is worth considering a dual
or even triple block. The PCR cyclers with this option allow to run up to 3 PCRs in parallel, all with different parameters. In this way, either the optimization can be simplified, or up to 3 people can carry out their PCRs at the same time. However, with many samples or the use of PCR plates, the networking of PCR cyclers is an
important additional function. Depending on the model, a main cycler can control up to 9 additional Eco-Cyclers.
And there's one more: When using control software on a desktop PC within a network, up to 50 Mastercycler X50 PCR Cyclers can be controlled individually. In this way, all cyclers can be in one or more rooms and can be conveniently operated from the desk. A message about the status of the device is visible at any time without running to it. This offers time to take care of paperwork and
relax while waiting for the end of the PCR. Such a program also offers additional security through user management and automatic storage of the performed PCR runs. If the CycleManager X50 software
is then connected to Visionize Labsuite, the audit trail is also secured and there is the option of connecting and checking other devices in the laboratory.
All about liquids
There are now a large number of PCR reagents on the market, including "Fast PCR Enzymes". These enable a much faster PCR than before using particularly fast polymerases. In this way, a PCR can be reduced from 40 minutes to 12 minutes. One could also say that 3 fast PCRs are carried out in the time of a classic PCR.
Another model is the implementation of multiplex PCRs
. Here, several amplicons of different sizes are amplified in just one PCR. And this method is also already available in the Fast PCR variant.
The right combination makes the difference
All in all, there are many optimization possibilities in the field of PCR, depending on what is needed. The fastest and most convenient option would be to use Fast PCR consumables and Fast PCR kits together with preparation in a pipetting station epMotion®
and a silver block cycler (Mastercycler X50s) with 2D gradient, controlled from a desktop PC with theCycleManager X50 software.
Another proposal: The work is quieter and more ergonomic due to the reduction in pipetting and physical distance to the devices.