Re: [DIYbio] DNA quantification instruments

Hello. As mentioned in a few previous posts I'm shopping for lab equipment, and one of the pieces I'm missing is some kind of equipment for quantifying DNA. I have considered several alternatives, and would like to ask about your experiences. To specify my needs a bit more, I am primarily interested in doing DNA concentration measurements of samples (PCR products) to be sent for sequencing. So preferably, the method should not require a too large sample volume.

Previously, I have used solely the NanoDrop (ND-1000) for measuring DNA concentrations. It's a nifty instrument and I'd love to have one but the prices I have seen have been in the $10 000 range, which is far above my current budget. Because it is a relatively new product, there are also few used units being sold. The main attraction of the NanoDrop is the microliter sample volume and ease of use. The main criticism I have seen of the ND, and which I agree with, is poor accuracy at low DNA concentrations with too high DNA concentrations being reported. However, the absorbtion curve being presented visually for each measurement at least makes it possible to evaluate how much trust to put in each measurement. There are several NanoDrop "clones" available too, such as Quawell (http://www.quawell.com/), but these are still priced at $5000 or so, and I'm unsure of the quality.

Although I'm unlikely to purchase one in the near future, I'd still like to hear your opinions on NanoDrop and other microliter-spectrometers. Are they worth the expense?

An instrument which I have seen touted as a cheaper and more accurate alternative to the NanoDrop is the Qubit Fluorometer.(http://www.lifetechnologies.com/no/en/home/brands/product-brand/qubit.html?cid=fl-qubit). Especially at low DNA concentrations, the Qubit seems to be more useful. The downsides are larger sample volume (up to about 20 uL needed), some sample preparation needed, and the need to buy instrument-specific consumables. Still, this instrument can be had for about 2-3000 $USD, which is affordable and would seem to give good value if the Qubit is as accurate as the advertisments suggests.

Have any of you used the Qubit or similar fluorescent-based DNA quantification instruments? What has your experience been?

The third alternative I am aware of is a plain old spectrometer. I also want something for measuring optical density of bacterial cultures, so a spectrometer is something I'd like or need for that purpose anyway. My main concern with using one for DNA concentration measurements is the need for (much) larger sample volumes than with NanoDrop or Qubit. Rather than buying a large, heavy and expensive all-round spectrometer, I am considering buying an Eppendorf Biophotometer (http://www.eppendorf.com/int/index.php?pb=63cfedee26795e18&action=products&contentid=1&catalognode=53772) (That's the newest model, but there are used older models for sale much cheaper). This seems to give a good all-round package for OD600, DNA/RNA and protein concentrations at smaller size and lower price. There are also small, cheap generalized spectrometers of Chinese make available, but I am very unsure of the quality of these...

Do you any of you have experience with the Eppendorf Biophotometer/Biospectrometer series or other spectrometers for measuring DNA concentrations? How much work goes into making standard curves and calibration? What is the minimum practical sample amount?

And finally, for those who have used several of the methods above: What do you recommend as the "go to" method for measuring DNA concentrations for a new DIYBio laboratory?

Looking forward to hear your opinions!

Best regards,

Jarle Pahr

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