it is monoplex not multiplex and not RFLP
Hi
Maybe contamination. Try to performance a new pcr with new products. What kind of tissue are you use?
Regards
Hi,
Its seem non specific amplification due to following reason:
(1) Nonspecific amplification due to too much template
(2) Many time I get same result when I amplify insert from plasmid having the insert due to low annealing temp.
(3) Use gradient PCR from annealing temp Tm-5 to Tm+5. many time using annealing temp more then Tm will surprisingly gives good specific amplification.
(4) Check primers for their binding specificity.
You can find a lot literature on PCR troubleshooting.
Hope this will help.
thnx
First my question what is the size of your product (how many base pairs).
Non specific amplification. Mention your PCR conditions and components.
As mentioned above try gradient. (If you have patience read my PCR answers I answered many).
Non specific and fragmented DNA. Clean up your DNA, increase annealing temp of primers and do titration of magnesium in your pcr mix
Non-specific amplification. I think if you increase magnesium concentration and annealing temperature, you can get rid of most of the background. Also, include positive and negative controls.
With my pleasure to you sir .
But I have not good information and poorly about this advance technique ?
take my respects .
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