Hindawi Publishing Corporation BioMed Research International Volume 2013, Article ID 270805, 4 pages http://dx.doi.org/10.1155/2013/270805 Research Article A Guide RNA Sequence Design Platform for the CRISPR/Cas9 System for Model Organism Genomes Ming Ma, 1 Adam Y. Ye, 2,3 Weiguo Zheng, 4 and Lei Kong 2 1 Biomedical Engineering Department, College of Engineering, Peking University, Beijing 100871, China 2 Center for Bioinformatics, State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, Peking University, Beijing 100871, China 3 National Institute of Biological Sciences, 7 Science Park Road, Zhongguancun Life Science Park, Beijing 102206, China 4 Institute of Computer Science and Technology, Peking University, Beijing 100871, China Correspondence should be addressed to Lei Kong; kongl@mail.cbi.pku.edu.cn Received 4 July 2013; Accepted 13 September 2013 Academic Editor: Yi Zhao Copyright © 2013 Ming Ma et al. his is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Cas9/CRISPR has been reported to eiciently induce targeted gene disruption and homologous recombination in both prokaryotic and eukaryotic cells. hus, we developed a Guide RNA Sequence Design Platform for the Cas9/CRISPR silencing system for model organisms. he platform is easy to use for gRNA design with input query sequences. It inds potential targets by PAM and ranks them according to factors including uniqueness, SNP, RNA secondary structure, and AT content. he platform allows users to upload and share their experimental results. In addition, most guide RNA sequences from published papers have been put into our database. 1. Introduction Gene engineering technology has always been a hot topic in life science research. With the development of gene modiication technology, certain genes can be knocked out or knocked down to a lower level. he appearance of zinc inger nuclease (ZFN) and tale nuclease (TALEN) has greatly accelerated progress in this ield, but their eiciency is oten unpredictable and it is diicult to target selected genes [1–8]. Recently, Cas9/CRISPR has been reported to successfully induce targeted gene disruption and homologous recombi- nation in both prokaryotic and eukaryotic cells with higher eiciency compared with ZFN and TALEN [9–13]. Addition- ally, it is easier to design guide sequence and easy to use for Cas9/CRISPR system [10]. his novel technology will be of great potential for application in both research ield and clinical trials. However, there is no available tool for the guide RNA design of Cas9/CRISPR silencing system. Although Mali et al. have reported the construction of unique whole human genome guide RNA library, covering more than 40% human exons [9], they did not provide a tool for researchers to design novel target sequences for other model organisms. Existed library also did not take into consideration related inluencing factors, such as SNP, deletion or insertion on the genome, and potential RNA secondary structure. According to our current understanding of the gRNA maturing process, the secondary structure of gRNA is crucial for Cas9-gRNA complex [14]. he 20 bp guide RNA sequence is used to bind with target site in genomes. If they are mostly involved into RNA loops, the eiciency to bind with target sites would be low. hus, this factor should be taken into consideration. Besides, the interference eiciency is probably closely related to the melting temperature of the gRNA-DNA hybrid. A relatively high AT content is negatively correlated with the of-target efect, and thus sequence with extremely low AT percentage is, to some extent, not recommended [9]. hus, we developed an online platform for the guide RNA design of the Cas9/CRISPR silencing system (http://cas9.cbi.pku.edu.cn/), with DNA variants information integrated. his tool helps researchers design their candidate guide RNA sequences more easily and provides assistance