Structural peculiarities of linear megaplasmid, pLMA1, from interfere with pyrosequencing reads assembly

Martin Wagenknecht (1) , Julián R. Dib (2) , Andrea Thürmer (3) , Rolf Daniel (3) , María E. Farías (2) , Friedhelm Meinhardt (1)

1 Institut für Molekulare Mikrobiologie und Biotechnologie
2 LIMLA - Laboratorio de Investigaciones Microbiologicas de Lagunas Andinas [Tucumán]
3 Göttingen Genomics Laboratory

Different strains of , isolated from high-altitude Argentinean wetlands, were recently reported to harbour the linear plasmids pLMA1, pLMH5 and pLMV7, all of which with 5′-covalently attached terminal proteins. The link between pLMA1 and the host's erythromycin resistance as well as further presumptive qualities prompted us to perform a detailed characterization. When the 454 technology was applied for direct sequencing of gel-purified pLMA1, assembly of the reads was impossible. However, combined Sanger/454 sequencing of cloned pLMA1 fragments, covering altogether 23 kb of the 110-kb spanning plasmid, allowed numerous sequence repeats of varying in lengths to be identified thus rendering an explanation for the above 454 assembly failure. A large number of putative transposase genes were identified as well. Furthermore, a region with five putative iteron sequences is possibly involved in pLMA1 replication.

Mots clés: 454 Sequencing; Inverted repeats; Iteron; Linear plasmid; repetitive sequences; Transposase

Martin Wagenknecht, Julián R. Dib, Andrea Thürmer, Rolf Daniel, María E. Farías, et al.. Structural peculiarities of linear megaplasmid, pLMA1, from interfere with pyrosequencing reads assembly. Biotechnology Letters, 2010, 32 (12), pp.1853-1862. ⟨10.1007/s10529-010-0357-y⟩. ⟨hal-00610056⟩