Purification of digested PCR product (a smear fr 200bp to 3kb) - (Aug/20/2007 )
Hi. I really hope that someone can help me here.
I digested a genomic DNA and ligated with a linker. Then PCR was carried out. The PCR product was a smear ranged from 200bp to 3 kb. Subsequently the linker was removed from the PCR product using RE. Next was to purify the digested PCR product from the cut linker because I need to ligate them with a second linker. I have tried many kits (PCR purification kit) and column (sephacryl 300HR prepacked column) but the recovery was poor. I need to generate 80 ug of purified PCR product. Can anyone HELP!!!
1) are you sure you need to do this? Wouldn't a new pair of PCR primers do the same job with much less trouble?
2) I'd switch to a loose silica technique, such as Geneclean, or more readily available and perhaps better, Qiaex II.