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Otherwise the quality of RNA is ok. But when I size select RNA with PAGE and elute RNA from the gel using 0.3M Sodium Acetate and precipitate with ethanol it results bad 260/230.
Lower the RNA concentration the worse 260/230 (can be in a range 0.2 - 2). The problem is increased increase 230 and 220 nm reading.
Wash (even repeated) of EtOH precipitate with 70% ethanol does not increase 260/230.
Initially the RNA is got with hot phenol treatment followed by ethanol precipitation (in the presence of sodium acetate). But even then in case of low amounts of RNA the 260/230 is good.
So seams that cause of bad 260/230 is arising from the gel it self (8% PAA 7M Urea TBE):
I noticed also:
1) second elution from gel gives better 260/230 for the same RNA concentration.
2) using NaCl instead of sodium acetate gives slightly better 260/230.
The question is what is this contaminant ruing 260/230 and how to get rid of it?
It should not be Urea as in my hands urea does not give similar 230 and 220 nm readings?
Could it be something from TBE or even acrylamide?
Lower the RNA concentration the worse 260/230 (can be in a range 0.2 - 2). The problem is increased increase 230 and 220 nm reading.
Wash (even repeated) of EtOH precipitate with 70% ethanol does not increase 260/230.
Initially the RNA is got with hot phenol treatment followed by ethanol precipitation (in the presence of sodium acetate). But even then in case of low amounts of RNA the 260/230 is good.
So seams that cause of bad 260/230 is arising from the gel it self (8% PAA 7M Urea TBE):
I noticed also:
1) second elution from gel gives better 260/230 for the same RNA concentration.
2) using NaCl instead of sodium acetate gives slightly better 260/230.
The question is what is this contaminant ruing 260/230 and how to get rid of it?
It should not be Urea as in my hands urea does not give similar 230 and 220 nm readings?
Could it be something from TBE or even acrylamide?