I prefer the CRISPR tool from the Zhang Lab at MIT. It gave me very good guides that worked nicely

http://crispr.mit.edu/

For a handy yeast (Saccharomyces cerevisiae) tool see:

http://yeastriction.tnw.tudelft.nl

A publication on this tool and its use has been accepted for publication in FEMS Yeast Research

there's lots of them, here's a non-exhaustive list i've managed to compile:

Thank you everyone for your replies, I really appreciate all the input. I hope to start working with this system in the next year and it is great to know that many of you have had success with these online tools.

Check out this following Addgene blog post, It has great comparison for all the gRNA designing software:

http://blog.addgene.org/the-crispr-software-matchmaker-a-new-tool-for-choosing-the-best-crispr-software-for-your-needs

I have tried out most of the CRISPR design tool listed in the Addgene blog post. I like CCTop (http://crispr.cos.uni-heidelberg.de/ ) best! The gRNA prediction it get is very similar to the Zhang Lab one (http://crispr.mit.edu/) but it tells you where the sequence locate (exon, intron, or non-coding area).

I suggest this tools. E-CRISP (http://www.e-crisp.org/E-CRISP/)

I suggest ADDGENE ( https://www.addgene.org/ ) where guide/help are being explained in details.

MIT CRISPR Tool

All these tools are very good, but I suggest to read the papers and design the gRNA manually. I have worked on designing some CRISPR vectors and gRNAs, and on different systems, and I figured out the best working gRNAs are the ones designed manually.

CHop CHop works - https://chopchop.cbu.uib.no/#

I pick sgRNAs from genome-wide libraries, this excel workbook can list and annotate the sgRNAs directly on the genbank sequence:

article:

https://www.linkedin.com/pulse/sgrna-annotator-excel-based-tool-pick-crispr-sgrnas-amir/

youtube:

https://www.youtube.com/watch?v=MO8rAcklmbI

github:

https://github.com/Ghahfarokhi/sgRNA_Annotator