How to improve DNA gel extraction process?

[Tim S]

Hey guys! 👋 

Does anybody know any tip to increase DNA yield from agarose gel? I use Promega Kit and short fragments clean up properly but when it comes to linear plasmid DNA (>5000 bp), yields are poor.

[StringJunky]

Is this any help. I put 'increase dna from agarose with promega'. This promega tutorial link came up.

https://www.promega.co.uk/resources/guides/nucleic-acid-analysis/dna-purification/

[Tim S]

Thank you for the answer! Looks very comprehensive. But the point is that, it doesn't work as the describe it unfortunately 

[Phi for All]

1 hour ago, Tim S said:

Thank you for the answer! Looks very comprehensive. But the point is that, it doesn't work as the describe it unfortunately 

"Looks comprehensive" is an admission that you didn't read it, which seems odd if you really want an answer. Should we assume you haven't contacted Promega about this problem with their product?

[Tim S]

I actually have read it very carefully. Thank you! By saying "very comprehensive" I meant I have never seen this long and meticulous article through I have read a lot of Promega protocols on this topic. 

Answering to your question- no, I have not contacted to Promega. I have read the text. And I mean their protocol specifics like "Persent Recovery" do not match to my experience of working with their Kits

[Phi for All]

37 minutes ago, Tim S said:

I actually have read it very carefully. Thank you! By saying "very comprehensive" I meant I have never seen this long and meticulous article through I have read a lot of Promega protocols on this topic. 

You're welcome! Perhaps we have different definitions of "very comprehensive", but I wouldn't describe an article that way if it didn't address my concerns over poor yields when using their product.

41 minutes ago, Tim S said:

Answering to your question- no, I have not contacted to Promega. I have read the text. And I mean their protocol specifics like "Persent Recovery" do not match to my experience of working with their Kits

Are you looking to add other voices to your complaint about their product before contacting them directly?

[Larmour Toujour]

I believe the silica spin columns for gel extraction are optimised for a certain DNA fragment size range. I can't remember whether the gel extraction kit columns are better for small or large fragments, but your could try using miniprep spin columns instead because these will be optimised for larger fragments.

[Larmour Toujour]

There are a few things you can do to improve the yield. 

1. Leave the DNA to bind the membrane for longer.

2. Elute using buffer that has been pre-warmed to ~50oC.

3. Pass the eluate through the column twice.

4. Elute in the smallest volume (15 uL).