DNAzol™ BD Reagent, for isolation of genomic DNA from whole blood

The color is due to cells lysing before the DNAzol Reagent is added. The color is caused by hemoglobin. This contaminant will cause problems during PCR, and must be removed before the ethanol wash step to prevent them. The following are possible reasons for the contamination:

- Inadequate or ineffective anticoagulants can result in clot formation; typically, this will give DNA a speckled appearance. Remove them by centrifugation before precipitating the DNA.
- The walls of the tube were not adequately washed with the DNAzol BD wash solution. Trace amounts of protein on the wall of the tube can cause contamination if this residue is dislodged during the ethanol wash steps.

The DNA isolated is actually total DNA, so plasmid DNA will be isolated along with genomic DNA. The mitochondrial genome is similar to a plasmid and can be isolated using DNAzol Reagent. The 1 minute room temperature incubation in ethanol before centrifugation should be extended to 5-10 minutes for maximum recovery.

We recommend storing DNAzol Reagent at room temperature. The DNAzol lysate (homogenate) can be stored 1 month at 15-30 degrees C; after 10 months at 4 degrees C or -20 degrees C, the DNAzol lysate (homogenate) has yielded high molecular weight genomic DNA, which can be completely digested with restriction enzymes and works well in PCR. During washes, DNA can be stored in 95% EtOH for at least one week at 15 degrees C to 30 degrees C or for three months at approximately 4 degrees C. DNA can be stored in DNAzol Reagent for one month at room temperature or 10 months at 4 degrees C.

Please see the following web page (http://www.thermofisher.com/us/en/home/life-science/dna-rna-purification-analysis/dna-extraction/genomic-dna-extraction/blood-dna-extraction.html) for a comparison of kits that can be used to isolate genomic DNA from blood.

Consider the following if you have a low 260/280 ratio:

The correct amount of DNAzol reagent may not have been used. If DNAzol reagent was added to a cell pellet, make sure that the volume of reagent is 20 times that of the cell pellet.

There may have been a problem in pipetting away the viscous supernatant from the DNA pellet, leading to contamination with protein. The DNA may be used with DNAzol reagent again or extracted with phenol to remove the protein.

In some samples dissolved in water, the ratio may be low due to the acidity of the water or the low ion content in the water. The ratios may go up if the sample is dissolved in TE and the spec is zeroed with TE (or 1 to 3 mM Na2PO4, pH ~8.0). [See BioTechniques 22:474-6, 478-81 (1997).]. The molar extinction coefficient of the nucleotides is given at neutral pH, suggesting that the absorbance at 260 nm would be highest at neutral pH. Of course, DNA is not stable under acidic conditions so degradation may occur if the DNA is left in this condition for too long.