I want to do UV-Melting for my G-quadruplex forming Oligonucleotides in presence of a specific metal ion. Our lab is new to this topic and no one in our lab has experienced about this.I have tried using some online protocol for UV melting sample preparation 2-3 times but it does not work good. I am also confused about which buffer to use so that neither its pH wont change with melting nor it affect G-quadruplex structure in anyway.Does anyone of you are using this techniques in your lab and if yes, can you please share the protocols for the same.I would be highly obliged.
My Colleague Richard Sheardy worked extensively in this area. His students' dissertations can be found at:
https://scholarship.shu.edu/do/search/?q=Sheardy&start=0&context=1927087&facet=
These will have the details you need
I worked on K+ stabilized G-quadruplexes. I suggest you three very good works that could be useful to prepare your samples, at least in Na+ and K+ environment. I followed these procedures myself. The articles are:
- Bianchi et al. 2018 Nucleic Acid Research - Structure of human telomere G-quadruplex in the presence of a model drug along the thermal unfolding pathway (look in Materials and Methods, that is the procedure I used in all my samples).
- Vorlickova et al. Methods 2012 - Circular dichroism and guanine quadruplexes (especially in chapter 15 you find protocols, tips and tricks. The preparations for CD and UV-Melting are the same).
- Renciuk et al. Nucleic Acid Research 2009 - Arrangements of human telomere DNA quadruplex in physiologically relevant K+ solutions. (this work extensively discuss about the differences in Na+ and K+ metal ions folding).
I hope this will be helpful
God luck.
This paper help you
Vorlickova et al. Methods 2012 - Circular dichroism and guanine quadruplexes (especially in chapter 15 you find protocols, tips and tricks. The preparations for CD and UV-Melting are the same).
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