Screening by tRNA primer extension analysis of porcine kidney mRNA libraries defines a novel endogenous porcine retroviral long terminal repeat

Academic Article

Abstract

  • Background. We have taken advantage of the common requirement of all eukaryotic retroelements for a specific tRNA primer to initiate DNA synthesis and applied a previously described in vitro screening methodology to the analysis of in vivo porcine tissues for transcriptionally active retroviral sequences. Methods. A series of 18-base pair (bp) 3' tRNA oligomers complementary to established primer binding sites for a variety of vertebrate retroviruses, retrotransposons, and retroposons were applied to primer extension analysis of kidney poly(A) mRNA. Primer extension products are predicted to represent 'strong stop' signals characteristic of the initial stages of retroviral transcription. Results. Several extension products were cloned, sequenced, and analyzed as probes for screening the porcine genome for potentially active retroviral sequences. We used this strategy to identify and clone a 655-bp 5' long terminal repeat of a porcine retrovirus with significant homology to the simian sarcoma virus. This transcriptionally active virus has an 82-bp U5 region, a conserved AATAAA polyadenylation sequence, a 39-bp repeat reminiscent of other retroviral enhancers, and a unique glycine primer binding site. Conclusion. Our results suggest that tRNA primer cloning can effectively identify novel retroviral elements.
  • Authors

    Published In

  • Transplantation  Journal
  • Digital Object Identifier (doi)

    Author List

  • Hoopes CW; Platt JL
  • Start Page

  • 1391
  • End Page

  • 1395
  • Volume

  • 67
  • Issue

  • 11