Hi,
Can anyone please highlight the difference between normal agarose and agarose low EEO? Any specification as to which to use for agarose gel electrophoresis?
Is it that extraction of pcr band from Agarose low EEO is much better than normal agarose?
Dear Haaris, usually the only difference between normal and Low EEO agarose is their SO4 anion contents. Low EEO agarose has negligible SO4 amount, so the movement of nucleic acid over the entire gel remains uniform without any retardation in their movement upon passing of current.
During the gel run, anionic groups of agarose are basically immovable components of the gel matrix, while the cationic counterpart of agarose migrates freely towards the cathode (-ve) and this results in low BOM (basis of mobility) and causes EEO (electroendosmosis), hence proper separation hinders. So for nucleic acids (like PCR products) low EEO agarose is preferable anyway.
Also, low EEO agarose has less background when is stained by ethidium bromide or Sybr Safe at postrun. For 0.2-23 kb nucleic acid fragments, low EEO agarose is the best option.
But for qPCR (real-time PCR) products 15-19% PAGE is the most suitable option, as the usual qPCR products are less than 150 bp and more often even less than 100 bp.
Regards....
I agree with Jawwad Ahmad and the agaroses low EEO, high EEO, medium EEO and special EEO differ mainly in their electroendoosmosis (EEO) values. These values are very important in the separation especially of proteins. All four agaroses can be mixed to meet the desired EEO value. This Agarose low EEO Standard has a very low EEO value and is recommended for the preparation of analytical and preparative gels with a very good resolution of nucleic acid fragments with sizes larger than 1000 bp (at higher concentrations even less than 1000 bp). It may be used at concentrations between 0.8 and 2 %. Agarose Low EEO has a very low DNA binding capacity and is suited for blotting experiments (Northern, Southern, ...), control of restriction digests etc. and can be used in all buffer systems.
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