2.2: Appendix II- Using the Denovix

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Nathan Reyna, Ruth Plymale, & Kristen Johnson
Ouachita Babtist University & University of New Hampshire

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How to Use the Denovix

Watch the YouTube videos regarding questions for the DeNovix.

(https://www.youtube.com/user/denovixvideos/videos)

The Denovix allows you to measure the concentration of DNA, RNA, and protein in a couple of microliters of the solution. It is best to use 1.5\(\mu L\) to 2\(\mu L\) for accurate measurements. It also has the ability to measure fluorescently labeled molecules.

Startup Denovix by touching the screen. Set up parameters for measuring nucleic acid/double stranded DNA (dsDNA).

Clean Denovix. Pipette 5-10 \(\mu L\) of water on the pedestal (metal part that was under the arm with a hole in the middle), close arm, and then wipe off water with a Kim-wipe(no paper towels).
You will now need to measure a blank. (This should be the same liquid your sample is dissolved in). Use the elution buffer you use in your Zymopure miniprep. Lift metal arm and pipette 2\(\mu L\) of your blank liquid onto the pedestal. (Avoid introducing bubbles.)

4. Gently lower arm and click on the blank button on the screen.

5. When complete, lift the arm and simply wipe clean with a Kim-wipe (do not use a paper towel!).
You are now ready to measure multiple samples.
(It is recommended you re-blank the instrument every 30 min.)

6. Add 1.5 or 2\(\mu L\) of your DNA sample and click measure on the screen.

Note: You do not have to clean with water between samples. Simply wipe the pedestal dry when you are ready for the next sample. Repeat for all samples.

7. Record DNA concentration (ng/\(\mu L\)), 260/280 and 260/230 ratios for all samples.
260/280 = a ratio of ~1.7- 1.80 is considered pure DNA (most important value)
260/230 ratios are commonly in the range of 2.0-2.2 (can vary a lot)

8. Clean Denovix. Pipette 10 \(\mu L\) of water on the pedestal, close arm, and then wipe off water with a
Kim-wipe….then close out the program. (Press home button.)

What do my values mean?

Link to DeNovix Website to learn more: https://www.denovix.com/tn-130-purity-ratios-explained/

260/280 Nucleic Acid Purity Ratios

“260/280 ratios are routinely used to determine the purity of nucleic acid measurements. This ratio is most commonly used to determine the presence of protein and or phenol in the isolated nucleic acid sample. Table 1 describes a general acceptable range for these ratios; however, they are not a guarantee of sample purity.”

260/230 Nucleic Acid Purity Ratios

“The 260/230 ratio is used to indicate the presence of unwanted organic compounds such as Trizol, phenol, Guanidine HCL and guanidine thiocyanate. Generally acceptable 260/230 ratios are in the range of 2.0 – 2.2. Values higher than this may indicate contamination with the aforementioned compounds.”

A 260/280 ratio below 1.5 does not render the DNA unsuitable for any application, but lower ratios indicate more protein contaminants are present which may interfere with downstream steps. 260/230 measure residual contaminants from the purification process. Low ratios may indicate impure DNA that will be less successful in downstream steps.