GENOME INFORMATION LABORATORY Identification and molecular characterization of actively expressed PERV LTR elements in Korean pigs Jae-Won Huh1, Hong-Seok Ha1, Dae-Soo Kim2, Byung-Wook Cho3, and Heui-Soo Kim1,2,* 1Division of Biological Sciences, College of Natural Sciences, Pusan National University, Busan 609-735, Republic of Korea 2PBBRC, Interdisciplinary Research Program of Bioinformatics, College of Natural Sciences, Pusan National University, Busan 609-735, Republic of Korea 3Department of Animal Science, College of Life Sciences, Pusan National University, Miryang 627-706, Republic of Korea http:// www.primate.or.kr Abstract Results & Discussion 1 Identification of actively expressed PERV LTR elements Porcine endogenous retroviruses (PERVs) in the pig genome could be infectious risk factors in xenotransplantation (pig-to-human). Long terminal repeats (LTRs) of PERV element have promoter activity which can affect the replication properties of single PERV and adjacent genes. Using the computational technique, we collected all pig genome information including genome and transcript sequences. Well conserved sequences in LTR sequences are used for the identification of actively expressed LTR elements Using the RT-PCR approach, three different pigs (wild, domestic, and hybrid with Yorkshire) are used for amplification and cloning. About one hundred individual clones were sequenced and analyzed. Multiple alignment indicated that different insertion and deletion (INDEL) and tandem repeated sequences are located within the LTR sequences. Transcriptional regulation of different subtypes of LTR elements are investigated by transient transfection assay, indicating that five different types of LTR elements are actively transcribed in pig genome. 2 Phylogenetic analysis of actively transcribed PERV-LTR elements and clone information Introduction 3 Transient transfection assay of PERV-LTR in different cell lines Materials & Methods Large White Korean domestic pig Korean wild pig RNA samples RT-PCR RT-PCR Analysis Sequencing Cloning References Subtype identification Cloning in Expression vector Luciferase assay Patience C, Takeuchi Y, Weiss RA (1997) Infection of human cells by an endogenous retrovirus of pigs. Nat Med 3: 276-282. Scheef G, Fischer N, Krach U, Tonjes RR (2001) The number of a U3 repeat box acting as an enhancer in long terminal repeats of polytropic replication-competent porcine endogenous retroviruses dynamically fluctuates during serial virus passages in human cells. J Virol 75: 6933-6940 GENOME INFORMATION LABORATORY