s421 t2 talens  (Addgene inc)

Journal: Cell Death & Disease

Article Title: pS421 huntingtin modulates mitochondrial phenotypes and confers neuroprotection in an HD hiPSC model

doi: 10.1038/s41419-020-02983-z

Figure Lengend Snippet: a TALEN-binding sequences flanking the S421 codon in HTT exon 9. b TALEN cleavage activity as assessed by Surveyor assay. c ssODN donor DNA with BglI restriction enzyme site for screening. d Verification of targeting and screening strategy in K562 cells. e Expected restriction fragment pattern with BglI screening. f HTT Exon 9 PCR followed by BglI screening identifies a number of targeted CAG180 HD hiPSC clones (black arrows); example shown is for S421(A) targeting; red arrows represent mixed clones. g Summary of targeting efficiency. h Phasing of targeted allele by allele-specific PCR followed by Sanger sequencing.

Article Snippet: Subsequently, S421-T1 and S421-T2 TALENs were cloned into pTAL-GFP (Addgene, #43858) and pTAL-RFP (Addgene, #43859) vector using BamHI and Xbal restriction enzymes, respectively.

Techniques: Binding Assay, Activity Assay, Clone Assay, Sequencing