Journal Article A high excision potential of TALENs for integrated DNA of HIV-based lentiviral vector.

Ebina, Hirotaka  ,  Kanemura, Yuka  ,  Misawa, Naoko  ,  Sakuma, Tetsushi  ,  Kobayashi, Tomoko  ,  Yamamoto, Takashi  ,  Koyanagi, Yoshio

10 ( 3 ) 2015-03-17 , Public Library of Science
DNA-editing technology has made it possible to rewrite genetic information in living cells. Human immunodeficiency virus (HIV) provirus, an integrated form of viral complementary DNA in host chromosomes, could be a potential target for this technology. We recently reported that HIV proviral DNA could be excised from the chromosomal DNA of HIV-based lentiviral DNA-transduced T cells after multiple introductions of a clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 endonuclease system targeting HIV long terminal repeats (LTR). Here, we generated a more efficient strategy that enables the excision of HIV proviral DNA using customized transcription activator-like effector nucleases (TALENs) targeting the same HIV LTR site. A single transfection of TALEN-encoding mRNA, prepared from in vitro transcription, resulted in more than 80% of lentiviral vector DNA being successfully removed from the T cell lines. Furthermore, we developed a lentiviral vector system that takes advantage of the efficient proviral excision with TALENs and permits the simple selection of gene-transduced and excised cells in T cell lines.

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