I want to do cloning , for that i need to get the whole gene which is 1.7 kb size. but im getting very faint bands in PCR.The DNA sample is from bacteria and is isolated by using kit. Im using around 100 ngs concentration of DNA for each reaction but also im getting very faint bands.
what could be the reason for this???????????
Sudhakar Pagal
Try different temperatures of annealing. As you are going to clone, I believe you are using a proofreading enzyme. Follow the reaction condition according to the manufacturer. If using Taq, the extension time is 1 min/kb. When using different annealing temperatures, include 55 deg also. Most primers anneal at this annealing temp.
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Tapiwa Nyakauru
Atiyeh Abdallah
is it possible to clone that faint band in the plasmid or you need a strong band for cloning. iam also experiencing the same problem
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