Proc Natl Acad Sci U S A. 1991 Aug 1;88(15):6711-5.

Stable transformation of Leptomonas seymouri by circular extrachromosomal elements.

Bellofatto V, Torres-Muñoz JE, Cross GA.

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Laboratory of Molecular Parasitology, Rockefeller University, New York, NY 10021.

Abstract

To define the cis-acting sequences necessary for gene expression and DNA replication in trypanosomatids, we have developed a selectable vector that can be grown in Escherichia coli and maintained stably in the insect trypanosomatid Leptomonas seymouri. The vector is relatively small (6 kilobase pairs) and contains a portion of the L. seymouri alpha-tubulin gene positioned in-frame with a truncated neomycin phosphotransferase gene that confers resistance to the aminoglycoside G418. This construct is maintained in cells as a high-copy-number circular extrachromosomal element containing several head-to-tail copies of the transforming plasmid. In L. seymouri, alpha-tubulin-neomycin phosphotransferase fusion RNAs are polyadenylylated and possess a trans-spliced mini-exon. Additional DNA sequences can be inserted into the vector, propagated, and expressed in transformed cells.

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Stable transformation of Leptomonas seymouri by circular extrachromosomal elements.

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