In our laboratory, we are currently utilizing the Opentrons Flex platform; however, we have encountered significant challenges with DNA purification using magnetic bead–based protocols. To address these limitations, we are exploring the possibility of integrating a vacuum manifold system to implement alternative and potentially more efficient DNA purification methods.
Is there someone that has already integrated such a system into the Opentrons Flex — or attempted something similar — for DNA purification?
Hi Vittoria, we are trying to optimise the DNA purification using beads as well - would you specify what challenges you ran into? It would be super helpful for us!
Hi Vittoria,
I’m also working on DNA purification workflows using magnetic beads. If you’re open to sharing more details about the specific challenges you’re encountering, I may be able to offer some suggestions based on my experience.
Hi everyone! We have spent several months trying to optimize the protocol, both on the robot and manually using the same kit. We ensured that the beads were perfectly resuspended and even increased incubation and elution times, as well as the frequency of resuspension, but results remain inconsistent.
The main issues we are facing are:
Low Yield: This occurs both manually and on the robot. However, after quantifying all waste fractions and the sample prior to adding the binding beads, we realized the DNA concentration is already low at the start, likely due to the initial inoculum volume. While low yield is an issue, it is not our primary concern as we can compensate for it by scaling up to 96 wells.
Purity: This is the most critical problem. On the robot, the eluate consistently comes out ‘dirty’ (contaminated). We attempted additional purification steps, but this led to an even greater loss of DNA.
Since the low yield can be managed by increasing the scale, our absolute priority is solving the purity issue while managing deck capacity. Currently, 24 samples occupy all available positions, including expansions. To reach 96 samples and maintain a fully automated workflow, we need to optimize tip usage and deck layout without compromising the quality of the final product.
Is this reagent contamination (GTC/salts), sample, or quality (260/280; 260/230)? And if sample contam, is it sourced from the process like cross contamination during extraction or from the source?
For most of these issues, I’d recommend trying a third low salt wash (or 80% EtOH) if your reagent reservoirs allow for extra volume.
Also relevant to the low yield: I’ve found in my own lab that extracting less cells per prep will yield higher overall yield - but at the expense of concentration. For instance, I found about 33% higher yield per cell when using 0.5MM cells per prep compared to 1.0MM cells per prep. We’ve optimized this to balance yield and concentration so we don’t need a subsequent concentration step, but definitely something to consider if you don’t mind playing with lower concentration eluate and highly value total yield. Helped us reduce our cell requirements for a non-PCR assay from 80MM cells to ~42MM cells, which was great.
For most extraction protocols, you’re going to consume 6-8 tips per sample unless you start re-using tips for reagents, waste removal, etc - it can get complicated. 24 samples sounds about right if you’re working with 9 or 12 worktable positions. You’ll have to empirically figure out if re-using tips is practical for your application otherwise you might be maxxed out.
I think at most I’ve reduced this down to 4 tips per sample, but that took quite a bit of applications time. This was also for plant samples… so cost was the most important factor to the end user of that protocol.
Hi Meghan, I’m in touch with someone from the company already but I am interested in knowing challenges other fellows have faced and how they overcome them!
This is an old post, but we encountered issues when we moved to a new DNA extraction bead based kit - and the problem was actually an interaction between our digest buffer (in house and) and the extraction reagents. A precipitate was being formed, which the beads got caught in, and then discarded with the waste. Just a thought …
@vittoria I second what @evwolfson said. We’ve gotten our protocol pretty consistent, and switching the final wash to 80% ethanol made a huge difference.