I did a genomic DNA extraction Gram negative bacteria by Wizard® Genomic DNA Purification Kit. Samples were treated with RNase Sigma-Aldrich, but spots appear at the end of the gel. I suppose it may be small RNA fragments. Lambda DNA/HindIII Marker. See figure below.
The conditions for the RNase treatment was with 15 uL concentration of 10 mg / ml for 60 minutes by 37 ° C.
How do I remove RNA in DNA extraction? I need pure high concentration of DNA.
It looks like you have good quality DNA. With the Qiagen kits, you add the RNase at the point where you resuspend the DNA pellet before cell lysis. This ensures that degraded RNA is removed during the column washes.You will need to clean up the DNA with a column. Alternatively you can perform a phenol/chloroform extraction followed by ethanol precipitation.
It happens some times that RNA remains in your DNA preparation even after RNase treatment, This happens may be due to improper RNase activity. RNase activity like any other enzyme activity is pH dependent. Make sure your DNA is dissolved in a solution with a pH favourable for RNase activity. Whatever residual RNA you are getting, that is because of prtial digestion due to sub-optimal activity of RNase. You can also incubate longer in RNase (may be 2 hours instead of 1) to ensure complete digestion before doing phenol-chloroform extraction.
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