Hi Sudip, I can answer your question partially due to this is out of my interest. Currently, the frequently used method for investigating of RNA-protein interaction is CLIP that stands for Cross-Linking ImmunoPrecipitation. But, you should know your target proteins that possibly interacting with ncRNAs to perform this method. Additionally, if you combine this method with next-gen sequencing (aka HITS-CLIP), you can also study transcriptome-wide interactions between the target protein and RNAs.

I am sending several links that may be helpful to you. 

http://epigenie.com/epigenetics-research-methods-and-technology/rnaprotein-interaction-analysis/rna-immunoprecipitation-rip-cross-linking-immunoprecipitation-clip/

http://www.ncbi.nlm.nih.gov/pubmed/25411354

http://www.ncbi.nlm.nih.gov/pubmed/24940765

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3961350/

Dear Volkan,

Thank you for the related information, this is really helpful. But for my RNA of interest we do not know any protein or RNA that interacts with it. This is what I am after.

Regards,

Sudip

Hi Sudip, I guess dChIRP method explained by Howard Y Chang group can be useful for your purpose. This technique is base on RNA and detects RNA-RNA, RNA-Protein & RNA-chromatin interactions.

http://www.nature.com/nbt/journal/v32/n9/full/nbt.2943.html

Good luck

Dear Sudip,

if you have really no idea of what your interaction partners may be you should consider tagging your sRNA with a MS2 tag. It's basically a stem loop structure added to your sRNA which can be specifically enriched by a MS2 coat protein fused to some protein you can select for, e.g. a protein you have a specifc antibody for or something like a maltose-binding protein, which can be purified on a Dextrin Sepharose column. Subsequently you should be able to perform proteomics or RNomics to screen for proteins/RNAs that interact with you candidate ncRNA. 

To see which targets your ncRNAs influences inside the cell you could also perform overexpression (e.g. pulse expression) or deletion if its intergenic. The most comfortable yet most expensive follow up experiments would again we proteomics (LC-MS/MS) or RNomics (RNA-seq).

I hope this helps you find a suitable technique for your purpose,

Good luck,

Philip

Hello Sudip,

At protein level, I think that the best thing you can do is follow Philip advise and use a MS2-GFP trap, it's a complicated protocol but gives good results.

As for detecting which mRNAs are the targets of this ncRNA you can use an assay of immunoprecipitation of AGO2. This molecule binds the ncRNA and the mRNA to silence. By overexpressing your ncRNA of interest you will be able to get more ncRNA-AGO2-mRNA and see which are the targets that you are looking for.

Luck!

Dear Philip,

Thank you so much for your well defined answer. I really like this idea. 

Best regards,

Sudip

Thank you Sudip for putting up this query. I was also having trouble finding ncRNA targets in malaria parasite. And now I have few strategies to dwell on.