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View additional product information for DNAzol™ Reagent, for isolation of genomic DNA from solid and liquid samples - FAQs (10503027)
10 product FAQs found
It is possible to see two phases after addition of ethanol if the amount of DNAzol Reagent was too low. Add more DNAzol Reagent and continue.
If the cells or tissue were washed with phosphate buffer solutions prior to DNA isolation, the phosphate may have been carried over and be inhibiting restriction enzymes. We recommend adding DNAzol Reagent to the DNA solution and reprecipitating with 0.5 volumes of 95% EtOH. Wash twice with 95%, dry briefly, and resuspend in 8 mM NaOH.
You can try incubating samples resuspended in 8 mM NaOH at 37 degrees C overnight to resuspend the DNA. You can also try incubating at 45 degrees C for 15 minutes.
Yes, this happens due to the dye in the reagent, and seems to be dependent on the volume of the stored reagent. The color change does not affect its performance.
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.
Trypsinization is not needed. Simply add 1 mL of DNAzol Reagent per 10 cm2 of the culture plate area. Rock the plate back and forth and pipet the contents into a tube for ethanol precipitation of genomic DNA. If you want to trypsinize to count your cells, pellet the cells that will be used for DNA isolation after trypsinization. Remove the supernatant completely by aspiration, then add DNAzol Reagent to the cell pellet. Pipet 3 to 4 times to lyse the cells completely before proceeding to the centrifugation step.
No, the 8 mM NaOH will not affect the DNA integrity. In fact, DNA is most stable at slightly alkaline pH (>7). You will find that the isolated DNA does not resuspend well in water and has even worse solubility in Tris buffer. (Water often has a pH of lower than 7 due to dissolved CO2 from the air. This slightly acidic water will actually cause degradation of your DNA!) The pH of the 8 mM NaOH is ~9 and can be easily adjusted with TE or HEPES once the DNA is in solution. (Over time, the solution becomes neutral upon exposure to air from dissolved carbon dioxide.)
No. Washing is not necessary.
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.