An in vitro transcription-based protocol for mapping HIV integration sites using lentiviral integration site sequencing

Identification of integration sites of human immunodeficiency virus (HIV) is crucial for developing antiretroviral strategies. Here, we present an in vitro transcription-based version of lentiviral integration site sequencing (LIS-seq), enabling rapid integration site mapping. We detail steps for the establishment of clonal cellular models, genomic DNA isolation and fragmentation, T7 in vitro transcription, poly(A) tailing, sequencing library generation, and the analytical pipeline. LIS-seq has been applied to clonal cellular models infected with an HIV-based vector. For complete details on the use and execution of this protocol, please refer to Więcek et al.¹.