CHOPCHOP: a CRISPR/Cas9 and TALEN web tool for genome editing

Abstract

Major advances in genome editing have recently been made possible with the development of the TALEN and CRISPR/Cas9 methods. The speed and ease of implementing these technologies has led to an explosion of mutant and transgenic organisms. A rate-limiting step in efficiently applying TALEN and CRISPR/Cas9 methods is the selection and design of targeting constructs. We have developed an online tool, CHOPCHOP (https://chopchop.rc.fas.harvard.edu), to expedite the design process. CHOPCHOP accepts a wide range of inputs (gene identifiers, genomic regions or pasted sequences) and provides an array of advanced options for target selection. It uses efficient sequence alignment algorithms to minimize search times, and rigorously predicts off-target binding of single-guide RNAs (sgRNAs) and TALENs. Each query produces an interactive visualization of the gene with candidate target sites displayed at their genomic positions and color-coded according to quality scores. In addition, for each possible target site, restriction sites and primer candidates are visualized, facilitating a streamlined pipeline of mutant generation and validation. The ease-of-use and speed of CHOPCHOP make it a valuable tool for genome engineering.

Figure 1.

Workflow of a CHOPCHOP CRISPR/Cas9 query. The home page of CHOPCHOP allows users to enter a gene name, genomic coordinates or a DNA sequence, and select an organism and TALEN or CRISPR/Cas9 mode. Advanced options can be toggled. The main results page presents the sgRNA or TALEN target sites within the gene architecture (exon, blue; intron, red), with each option color-coded according to ranking. Hovering over an entry in the table highlights the corresponding graphical sgRNA/TALEN and vice versa. Clicking on a specific result takes the user to a page containing the zoomed in locus with the predicted cut site highlighted in red, primer options in purple and restriction sites color-coded according to whether they are unique in the region.

Figure 2.

CHOPCHOP provides detailed information about each CRISPR/Cas9 and TALEN target site. (A) The detailed information page provides a zoomed in view of the target locus with visible DNA sequence, primer options (above the gene, purple) and restriction sites (below the gene; green if unique in the region, red if not). In TALEN mode the target site is color-coded; TALEN 1 is blue, the spacer is black and TALEN 2 is red. (B) Information is provided about predicted off-targets: the genomic location, number of mismatches and location of mismatches within the sequence. (C) Information is provided about the primer designs, including the primer sequence, T_m and product size.