how to deal with PCR inhibitors ? what should we add or do to remove PCR inhibitors...
Perform spin column based isolation of DNA, while doing so dont exceed the rpm in centrifugation steps recommended by manufacturer. There are many kits for animal and plant dna isolation based on spin column. Prefer qiagen
Try to dilute DNA template (to such as 1/10x, 1/50x or 1/100x,.....) and do PCR amplification again to see whether it works.
Alternatively, re-purify the DNA sample, and check the purity with OD260/OD280 ratio.
Several methods are employed to remove PCR inhibitors depending on the what kind of sample you are subjecting for PCR. Based on your sample, use a specfic method to remove it. this review article on PCR inhibitors may help you.
In addition to the dilution of template, incorporation of formamide and DMSO in PCR reaction remains handy to address PCR inhibitors.
There are many ways in which you can handle PCR inhibitors, depending on the source and kind of inhibitor. I suggest reading the attached paper as a starting point. Hope it helps.
Article The peril of PCR inhibitors in environmental samples: the ca...
Hi
The first step is to obtain DNA in it's highest level of purity. For that you can use the phenol-method that will extract your DNA in an almost easy way and with a high degree of purity, since the phenol will desaturate everything that is not nucleic-acid. If you do the correct protocol, and with the correct periods of centrifugation, you will get DNA with a high degree of purity - which won't interfere with your PCR experiment.
But be sure to not use too much DNA per PCR sample, remember that the DNA-polymerase does is inhibit by high concentrations of DNA. Also, use the correct amount of co-factor ion and so on.
All this will contribute for good PCR experiment.
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