Nanodrop is just detecting the absorbance, and cannot distinguish DNA from dNTP.

Your PCR reaction contains lots of dNTP and primers.  Your just detecting the absorbance of them.

Nanodrop is just detecting the absorbance, and cannot distinguish DNA from dNTP.

Your PCR reaction contains lots of dNTP and primers.  Your just detecting the absorbance of them.

Hi Joah,

I agree with Artur!

have a look for this link for possible reasons for no bands. Good luck

Ali

https://www.fws.gov/aah/pdf/pcrts1_noband.pdf

Did you have dimer-primer?if so then you have to lessen the concentration of the primer in reagent or change the annealing temperature 

Reasons can be a lot:

1. badly designed primers

2. poor quality of temple (nucleic acid degradation)

3. inhibitors of the PCR reaction (contamination of reagents)

4. no expression or low expression of interest gene, (too small amount of template)

5. polymerase (Pfu polymerase is more effectively)

6. difficult temple (high GC content) (you need a kit for difficult temples)

7.  badly design PCR temperature profile (perform: gradient PCR or touchdown PCR)

Real time PCR it is a more sensitive method. A good idea is to prepare melt curves. If you have one single peak on melt curve its mean that you have one product. 

If you do not prepare allele-specific PCR, try to design new primers in another place of temple. More precisely, check the parameters of primers already in silico.

Good luck!

when check DNA concentration with nanodrop, A260/A280 and A260/A230 is important to determine whether the DNA is pure or not. if the ratio is too low or too high, the concentration value is not correct.

Thank you guys! It's been a while since I did cloning so the PCR purification step was forgotten...it's been now resolved :)