Do you have your target molecule coupled in the beads? And also what elution buffer are you using? For example I do the aptamer elution at 80°C and I haven't had any problems before
Hi,
Thank you for your reply.
Yes my target molecule is coupled beads.
My elution buffer is :
MgCl2, NaCL, Urea and tris -Hcl.
For me the color changed to Brown, and when I finish my selex and did cloning I had bleu and White colonies but after my sequencing my results were weard!!!
Can you help me plz.
Thank you
Siham
Hey,
It's quite strange this happens to you as this are still normal solutions used in SELEX protocols. Are the beads you are using bought recently? Perhaps if you had them a long time they are not as good any more
The elution buffer I'm using only contains Tris-HCl, Urea and EDTA but still NaCl and MgCl2 are used during the selection process anyway so it doesn't make sense that this is happening.
You could try to incubate the same solution with your beads in lower temperatures (e.g. 80 °C ). I still think 90 °C is quite high! Urea degrades in high temperatures so that could be a reason.
If the same happens again then perhaps you could try a different elution buffer. Have you taken this elution buffer from a specific paper? Look up papers that are using Tris, EDTA, Urea elution buffers instead and maybe give that a try.
Hope this helps
Hi;
Thank you for your replay. I bought the immobilization beads about two years ago. How can I know the efficiency of the beads!!Foor color changing I had it when my beads are new also!! I dont know what can I do!!! For the protocol I took it for different papers.
When you used Tris, EDTA, Urea elution buffers you heat or no! can I have the protocol for this!
Thank you.
Siham
Yes I heat the sample at 80 °C!
Have a look at the FluMag SELEX paper from Stoltenburg for the elution process
- Badges
- Science method