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Unexpected Instability of Family of Repeats (FR), the Critical cis-Acting Sequence Required for EBV Latent Infection, in EBV-BAC Systems

2011; Public Library of Science; Volume: 6; Issue: 11 Linguagem: Inglês

10.1371/journal.pone.0027758

1932-6203

Teru Kanda, Sachiko Shibata, Satoru Saito, Takayuki Murata, Hiroki Isomura, Hironori Yoshiyama, Kenzo Takada, Tatsuya Tsurumi,

Plant Virus Research Studies

A group of repetitive sequences, known as the Family of Repeats (FR), is a critical cis-acting sequence required for EBV latent infection. The FR sequences are heterogeneous among EBV strains, and they are sometimes subject to partial deletion when subcloned in E. coli-based cloning vectors. However, the FR stability in EBV-BAC (bacterial artificial chromosome) system has never been investigated. We found that the full length FR of the Akata strain EBV was not stably maintained in a BAC vector. By contrast, newly obtained BAC clones of the B95-8 strain of EBV stably maintained the full length FR during recombinant virus production and B-cell transformation. Investigation of primary DNA sequences of Akata-derived EBV-BAC clones indicates that the FR instability is most likely due to a putative secondary structure of the FR region. We conclude that the FR instability in EBV-BAC clones can be a pitfall in E. coli-mediated EBV genetics.