Embryonic expression of endogenous retroviral RNAs in somatic tissues adjacent to the Oikopleura germline.
Bottom Line: Tor Env proteins are membrane-associated glycoproteins which exhibit some features of viral membrane fusion proteins.Such embryonic expression depends on determinants present in the Tor elements and not on their surrounding genomic environment.Our study shows that unusual modes of transcription and expression close to the germline may contribute to the proliferation of Tor elements.
Affiliation: Sars International Centre for Marine Molecular Biology, University of Bergen, Bergen, N-5008, Norway email@example.com.Show MeSH
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Mentions: For the synthesis of RNA probes (Supplementary Figure S1), we amplified gene fragments with polymerase chain reaction (PCR) using specific primers and cDNA libraries from mixed developmental stages (1–6 h pf). The PCR products were cloned in pCRII-TOPO (Invitrogen) or pGEM-T vectors (Promega), prior to linearization for in vitro transcription with T7 or SP6 RNA polymerases (Roche). For expression in HEK293T cells, we cloned the complete env ORFs in C-terminal fusions with V5/6His tag in pCDNA 3.1-TOPO vector (Invitrogen). To construct pCTor3b-2 and pCTor4b-1, we generated DNA fragments containing part of env, fused with the V5/6His tag, followed by a stop codon and a restriction site (Figure 6A). These were then digested with the appropriate enzymes and ligated to upstream and downstream DNA fragments in order to reconstruct a Tor element that carried the full gag to the 3′LTR sequence. The resulting inserts were PCR amplified and cloned into the pCRII-TOPO vector.
Affiliation: Sars International Centre for Marine Molecular Biology, University of Bergen, Bergen, N-5008, Norway firstname.lastname@example.org.