Can anyone help regarding QIAEX II Gel Extraction Kit?

An efficient purification and fractionation of genomic DNA from soil by  modified troughing method

What to do when degrated genomic DNA is the only option for Fundamentals Explained

Otherwise, you run the risk of that the DNA exposed to UV for the longest time will be nicked to shreds. Additionally, utilize a noticeable range stain such as methylene blue or crystal violet.  Did you see this?  can learn more about this step in our article on preparing vectors for cloning. 3. Get rid of All Traces of Phenol Using A "Home Brew" Approach, If you are using phenol to cleanse the DNA from agarose, bear in mind that phenol traces will not be eliminated by ethanol precipitation and will hinder subsequent ligation responses.

Then, let it cool back down to space temperature over 10-20 minutes before precipitating to guarantee that you get double-stranded DNA (keep in mind that double-stranded DNA separates a high temperature levels). 4. Change to a New Brand Name or Bottle of Agarose, Often, agarose actually causes enzyme inhibition during downstream reactions (e. g.

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It might be that the agarose is old and the quality is no longer excellent or it may brand-dependent. We can't offer a concrete reason for this observation, but bear in mind that merely changing to a different bottle of agarose may increase your chances of cloning success. 5. Run Controls, To figure out if your issue is really connected to the gel extraction procedure, try running a control in which you absorb empty vector with a single enzyme, perform the gel extraction, and re-ligate it.

The Greatest Guide To Zymoclean™ Gel DNA Recovery Kit w/ Zymo-Spin™ I

If the control ligation works, then the failure to clone your DNA construct might be related to some other element, such as secondary structure of the DNA, repeat series triggering instability in E.coli, or the DNA might encode a protein that is harmful in bacteria. The Following Tips Apply If You Are Using Industrial Silica Spin Kits:6.

This combination will denature the DNA. If the eluted DNA appears at half the anticipated size (it is now single-stranded), renature the DNA by warming it as much as 95C for 1 minute and let it gradually cool down to room temperature level. 7. Wash It Once again, An extra cleaning action with the ethanol-containing wash buffer in the kit will constantly assist get rid of chaotropic salt residues on the membrane.