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

WAGENKNECHT, MARTIN; DIB, JULIÁN RAFAEL; THÜRNER, ANDREA; DANIEL, ROLF; FARÍAS, MARIA EUGENIA; MEINHARDT, FRIEDHELM

BIOTECHNOLOGY LETTERS

SPRINGER

Año: 2010

0141-5492

Different strains of Micrococcus luteus, isolated from high-altitude Argentinean wetlands were previously found to harbor the linear plasmids pLMA1, pLMH5, and pLMV7, respectively, all of which with 5’ covalently attached terminal proteins. The recently reported link between pLMA1 and the host’s erythromycin resistance as well as further presumptive features implicated in other qualities prompted us to perform a more detailed characterization of the plasmid. When the 454 technology was applied for direct sequencing of gel-purified pLMA1, assembly of the reads encountered unexpected difficulties. Eventually, combined Sanger/454 sequencing of cloned pLMA1 fragments, spanning althogether 23 kb of 110 kb entire plasmid size, allowed us to identify numerous sequence repeats of varying length, concomitantly rendering an explanation for the above obstacles, as well as a large number of putative transposase encoding genes; furthermore, a region, exhibiting five putative iteron sequences, possibly involved in pLMA1 replication, was found.