Dear Shima,
I think you should examine PCR troubleshooting book that could find anywhere. If you get only weak band, you should increase your DNA amount or cycles number. If you have smear, you should calculate your DNA purity and you should get more pure DNA extraction. And more of them, when you examine troubleshooting book, get more answer for your question.
Old PCR primers aliquote that you were using might be one of the reasons of getting faint bands. Because, repeated freeze and thaw of primers degrades the quality of primers. And the second most common reason is the template DNA. If possible get a fresh template DNA and go with the same PCR conditions. Good luck.
If the primers were dissolved in water not TE then acid depurination of the oligos from CO2 absorption from the air may have degraded the primers. Order new primers and dissolve in TE
Hi @Andrew Octavian Sasmita,
Thank you so much for sharing the links. Do you happen to have the part II of the link below? I tried to find it online but to no avail. Is it available yet? Thank you very much, cheers.
- https://www.fws.gov/aah/PDF/PCRTS1_NoBand.pdf
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