One of the technique to amplify specific band of PCR product is gel touch tip but what I get is always smear effect as seen in gel pic below, even gel Band extracted and filtrateor elution used as PCR template still i do get smear effect. I did tried to purify by phenol :chloroform , still the results are same what i am doing wrong ?
Dear Priti Khatri
plz. check
https://www.researchgate.net/topic/Agarose-Gel
Hii,
I think your issue is DNA purification. Did you check the sample under nanodrop for purity? I could suggest you better if you share nanodrop values along with 260/280 and 260/230 values.
Though I can speculate 2 possibilities you can go check it back.
1. It may occur by RNA contamination, RNAse treatment can sort this problem.
2. make sure your solvent is absolutely nuclease free water.
Hope this will work
Regards
Dinesh
Your pcr product is getting sheared even after using gel band extracted DNA as template. So, running a gel with gel band DNA after gel extraction may be helpful to check integrity of template. I hope you have used DEPC treated or nuclease free water in all required steps during gel running, otherwise, you could follow it.
Best wishes...
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