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J Virol, April 1998, p. 2663-2670, Vol. 72, No. 4
0022-538X/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

A Replication-Competent Retrovirus Arising from a Split-Function Packaging Cell Line Was Generated by Recombination Events between the Vector, One of the Packaging Constructs, and Endogenous Retroviral Sequences

Heung Chong,1 William Starkey,2 and Richard G. Vile3,*

Division of Histopathology, United Medical and Dental Schools of Guy's and St. Thomas's Hospitals,1 and Department of Virology, St. Thomas' Hospital,2 London SE1 7EH, and Laboratory of Molecular Therapy, Imperial Cancer Research Fund Molecular Oncology Unit, Hammersmith Hospital, London W12 0NN,3 United Kingdom

Received 29 August 1997/Accepted 22 December 1997

Previously we reported the presence of a replication-competent retrovirus in supernatant from a vector-producing line derived from a widely used split-function amphotropic packaging cell line. Rigorous routine screening of all retroviral stocks produced in our laboratory has not, previously or since, indicated the presence of such a virus. Replication-competent retroviruses have never previously been used in our laboratory, and stringent screening of all routinely used cell lines has not revealed the presence of any helper viruses. Therefore, it is highly unlikely that this virus represents an adventitious cross-contaminant or had been imported unknowingly with our cell line stocks. PCR studies with DNA from infected cell lines and Northern blot analysis and reverse transcriptase PCR with RNA from infected cells suggest that the helper virus arose by recombination events, at sites of partial homology, between sequences in the vector, one of the packaging constructs, and endogenous retroviral elements. These recombinations were not present in stocks of the packaging cell line or in an initial stock of the vector-producing line, indicating that these events occurred while the vector-producing line was being passaged for harvest of supernatant stocks.

* Corresponding author. Mailing address: Laboratory of Molecular Therapy, ICRF Molecular Oncology Unit, Hammersmith Hospital, Du Cane Rd., London W12 0NN, United Kingdom. Phone: 44 181 383 3257. Fax: 44 181 383 3258. E-mail: R.Vile{at}icrf.icnet.uk.




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