The ability of the novel NEXTGENPCR thermocycler to perform ultrafast cycling often prompts questions from scientists. Here are the top ten questions and answers.
No. It is possible to get a nice gain in speed using your current PCR setup. However, with some optimization, which could include using an enzyme with an increased base per second incorporation speed, you can maximize the speed. This is useful when doing repeated PCR of the same target such as in screens or other high-throughput applications.
The microplates used in NEXTGENPCR are specific to the instrument as they have a wall thickness of 40 microns, five times thinner than the usual thin-walled PCR tubes. This design allows the slight compression of the blocks in each thermal zone to immediately bring the sample to the desired temperature while also mixing the sample.
Since the plates have a structural frame and match the SBS microplate layout, you can incorporate robotics into the workflow for loading the samples and placing the plates into the thermocycler. Anvils are available for the various well configurations that support the wells and prevent piercing the thin polypropylene during loading and retrieval of samples.
No. Amplification of a 100 bp fragment was optimized to less than two minutes. As expected, larger fragments will take longer. However, with optimization, we amplified a 1 kb fragment in only ten minutes. The time saved will depend on the fragment size and level of optimization done, but even using your current protocol, you can save time by eliminating ramp times. Just a little optimization can reduce the time further. By switching to a fast enzyme, you can achieve an additional gain in speed. The amount of optimization all depends on your needs.
You can use NEXTGENPCR to prepare samples for NGS. To achieve maximum speed, some protocols may have to be optimized or altered from their recommended kit protocol.
The NEXTGENPCR thermocycler has a small footprint such that it fits easily on the lab bench and still leaves room to work. The thermocycler measures 45 x 20 x 29.5 cm (l x w x h) and weighs 18 kg.
The unique plates must be heat sealed. A separate heat sealer is required to run plates on NEXTGENPCR. We offer a heat sealer; however, a specific heat sealer is not required to operate the instrument. Regardless of the heat sealer used, a NEXTGENPCR heat-sealing anvil is required to tightly seal each plate well. Pipetting anvils are available for the various well configurations to support the wells and prevent piercing the thin polypropylene during loading and retrieval of samples. Once the plate is sealed, you cannot remove the seal, and the sample is retrieved by puncturing the seal with a pipette tip. Piercing tips are available but not required to retrieve the sample; in most cases, a pipette tip available in your lab can provide the same result.
There is a USB port that is used for doing software updates.
The plates are the same size as a typical microplate and have the same outside rim. They adhere to the standard defined by the Society for Laboratory Automation and Screening, so they fit in all robotics – both upstream and downstream. However, they differ in the center where polypropylene film is contained that has either 96 or 384 wells with a well thickness of 40 microns. PCR mix is pipetted into the wells and then sealed using heat.
It is possible to perform reverse-transcription PCR on the instrument. However, the current software requires the user to program two separate protocols. The instrument is end-point PCR only and does not perform qPCR.