RT Journal Article SR Electronic T1 Scalable Design of paired CRISPR Guide RNAs for Genomic Deletion JF bioRxiv FD Cold Spring Harbor Laboratory SP 052795 DO 10.1101/052795 A1 Carlos Pulido-Quetglas A1 Estel Aparicio-Prat A1 Carme Arnan A1 Taisia Polidori A1 Toni Hermoso A1 Emilio Palumbo A1 Julia Ponomarenko A1 Roderic Guigo A1 Rory Johnson YR 2016 UL http://biorxiv.org/content/early/2016/05/22/052795.abstract AB Using CRISPR/Cas9, diverse genomic elements may be studied in their endogenous context. Pairs of single guide RNAs (sgRNAs) are used to delete regulatory elements and small RNA genes, while longer RNAs can be silenced through promoter deletion. We here present CRISPETa, a bioinformatic pipeline for flexible and scalable paired sgRNA design based on an empirical scoring model. Multiple sgRNA pairs are returned for each target. Any number of targets can be analyzed in parallel, making CRISPETa equally appropriate for studies of individual elements, or complex library screens. Fast run-times are achieved using a precomputed off-target database. sgRNA pair designs are output in a convenient format for visualisation and oligonucleotide ordering. We present a series of pre-designed, high-coverage library designs for entire classes of non-coding elements in human, mouse, zebrafish, Drosophila and C. elegans. Using an improved version of the DECKO deletion vector, together with a quantitative deletion assay, we test CRISPETa designs by deleting an enhancer and exonic fragment of the MALAT1 oncogene. These achieve efficiencies of ≥50%, resulting in production of mutant RNA. CRISPETa will be useful for researchers seeking to harness CRISPR for targeted genomic deletion, in a variety of model organisms, from single-target to high-throughput scales.