If the goal is to obtain DNA of maximal size, shaking during lysis in the thermal mixer is not required. However, Proteinase K digestion needs to be carried out at 56°C. To ensure proper lysis, invert your sample a number of times slowly during the proteinase K incubation. Please note that samples lysed without agitation will be very viscous and difficult to handle. If the goal is DNA tuned to 50 kb-250 kb (optimal for ligation-based nanopore sequencing), then we strongly recommend working with a thermal mixer (e.g., Eppendorf® ThermoMixer® C). This will result in a controlled shearing to the optimal fragment size range. If access to a thermal mixer is not possible, the best alternative for agitation in the thermal mixer is to pulse-vortex for one minute at the highest speed possible after the lysis incubation. For tissue samples, briefly vortex the sample every minute or so during the beginning of the lysis incubation, until there are no tissue pieces remaining. Then, vortex again for one minute after the lysis incubation.
Find a Golden Butterfly for a Chance to Win!
Help us celebrate our 50th anniversary! We have hidden 1,000 golden butterflies and are waiting for you to find them. They can be anywhere that you find NEB! Beginning April 15th, be sure to visit our website and tables at tradeshows and events you are attending. Visit our social media channels frequently for tips on where we have hidden the butterflies – and once you find one, either click or scan the code to be eligible for a 50th anniversary prize pack, as well as a grand prize trip to NEB headquarters in Ipswich, MA!.
