SAPTA: a new design tool for improving TALE nuclease activity

Abstract

Transcription activator-like effector nucleases (TALENs) have become a powerful tool for genome editing due to the simple code linking the amino acid sequences of their DNA-binding domains to TALEN nucleotide targets. While the initial TALEN-design guidelines are very useful, user-friendly tools defining optimal TALEN designs for robust genome editing need to be developed. Here we evaluated existing guidelines and developed new design guidelines for TALENs based on 205 TALENs tested, and established the scoring algorithm for predicting TALEN activity (SAPTA) as a new online design tool. For any input gene of interest, SAPTA gives a ranked list of potential TALEN target sites, facilitating the selection of optimal TALEN pairs based on predicted activity. SAPTA-based TALEN designs increased the average intracellular TALEN monomer activity by >3-fold, and resulted in an average endogenous gene-modification frequency of 39% for TALENs containing the repeat variable di-residue NK that favors specificity rather than activity. It is expected that SAPTA will become a useful and flexible tool for designing highly active TALENs for genome-editing applications. SAPTA can be accessed via the website at http://baolab.bme.gatech.edu/Research/BioinformaticTools/TAL_targeter.html.

Engineering synthetic TALE and CRISPR/Cas9 transcription factors for regulating gene expression

“…In fact, ongoing bioinformatics efforts are leading to improved algorithms for both designing TALENs with high activity (105) and specificity (106). Nonetheless, we and others have already found TALEs to be a very effective platform for designing custom enzymes.…”

Quantifying on- and off-target genome editing

“…TALENs can be engineered using multiple different methods, which usually require 1–2 weeks of relatively sophisticated molecular biology. In general, ~70% of engineered TALEN pairs show reasonable on-target activity when designed using appropriate tools [25], and the platform generally has greater specificity than ZFNs. Finally, the RGEN’s are the simplest to engineer requiring only basic molecular biology skills with a success rate of 30–66% [26].…”

Cellular Therapies: Gene Editing and Next-Gen CAR T Cells

“…TALEN target sites are readily found and TALENs are easier to design than ZFNs due the straightforward pairing of DNA sequence and RVDs (Cermak et al 2011 ). Guanines are the exception as they are commonly targeted using either using the RVDs NN or using NK, which provides more specifi city, though may lead to less activity (Streubel et al 2012 ;Lin et al 2014a ). A number of solutions of different scale were presented that alleviated the diffi culty of cloning the large, highly repetitive TAL domains (Cermak et al 2011 ;Zhang et al 2011 ;Weber et al 2011 ;Reyon et al 2012 ).…”