Results: 3

1.
Fig. 1

Fig. 1. Schematic diagram of the strategy for constructing TALENs via Golden Gate cloning method. From: Rapid and Cost-Effective Gene Targeting in Rat Embryonic Stem Cells by TALENs.

A: the structure of plasmid pTAL3 and pCAG-TAL3. B: the TALEN target is located in exon 12 of BMPR2 with a Hind III site in the spacer region (red). The left and right TALENs binding sequences are underlined. The pCAG-BMPR2-TAL-F was constructed using two rounds of Golden Gate cloning.

Chang Tong, et al. J Genet Genomics. ;39(6):10.1016/j.jgg.2012.04.004.
2.
Fig. 3

Fig. 3. BMPR2 gene disruption in rat ES cells mediated by TALENs. From: Rapid and Cost-Effective Gene Targeting in Rat Embryonic Stem Cells by TALENs.

A: PCR screening of the rat ES cell colonies. The positive colonies yielded an extra band on the gel, indicated by the arrow. B: DNA sequencing result of the extra band indicated in (A). C: alignment of the sequencing result with the intact BMPR2 sequence indicates a 104 bp deletion (highlighted). D: BMPR2-targeted rat ES cells stained for pluripotency markers Oct4 (top panel) and Nanog (bottom panel). Left panel: phase contrast images; right panel: nuclear counterstain with diamidino-2-phenylindole (DAPI).

Chang Tong, et al. J Genet Genomics. ;39(6):10.1016/j.jgg.2012.04.004.
3.
Fig. 2

Fig. 2. Confirmation of BMPR2 gene disruption in HCC cells. From: Rapid and Cost-Effective Gene Targeting in Rat Embryonic Stem Cells by TALENs.

A: the PCR strategy for identification of targeting events. As indicated, the primer pair F and R were used to amplify the 506 bp DNA fragment including the TALENs target site. B: the products from samples transfected with TALENs-expressing plasmids were resistant to Hind III digestion (lane 1), whereas non-transfected sample products were completely digested (lane 2). Untreated PCR products were loaded as a control (lane 3 and lane4). M1, 100 bp DNA ladder; M2, 1 kb DNA ladder. C: DNA sequencing results indicated a 9 bp deletion in the spacer region. The two TALEN binding sites are highlighted.

Chang Tong, et al. J Genet Genomics. ;39(6):10.1016/j.jgg.2012.04.004.

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