Hello Everyone,
the description of NEBuilder HiFi DNA Assembly mix says: "No PCR clean-up step required".
I always used this mix after DNA extraction from agarose gel , but now we have some problems with this step... Does anyone use this kit without PCR clean up, directly on the PCR product? Does it work? How many of the product is necessary? How could I measure or estimate the concentration of the DNA in this case?
Thank you very much,
Magdi
Hi Magdolna Keszthelyi , I 've nerver used the NEb HIFI without the purification on agarose before but if you want to try I think you can estimate your concentration of DNA on agarose gel with the DNA ladder you're using. You can load a few microliters and estimate (approximately) the concentration with the intensity of the band you obtain (you can read about it here https://www.researchgate.net/post/How-do-I-quantify-the-DNA-accurately-on-agarose-gel);
May I ask what problem you're having with the purification on gel ?
Have a nice day
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