Archaeal rDNA can be retrieved using eukaryotic specific primers

COVACEVICH F; CUMINO A; SILVA R; NEGRI R; SALERNO GL

Quilmes, Buenos Aires

Congreso; Tercer Congreso Argentino de Microbiología General; 2006

Sociedad Argentina de Microbiología General SAMIGE

The aim of this work was to recovery picoeukaryotic organisms from the environmental samples of the Argentinean Sea. Picoplankton were collected from surface well-mixed water collected in September, October and November 2003 at the fixed EPEA Station (Mar del Plata, Argentine). Nucleic acid (DNA) extraction was performed, using the phenol-chloroform method. Extracted DNA was used as the template in a PCR reaction using eukaryotic 18S ribosomal DNA (rDNA)-specific primers. Two bands were obtained at the agarose gel electrophoresis from the samples extracted in September and October. Higher bands were about 1700 kb and lower bands were about 1500 kb. Only one band of about 1700 kb was obtained from sample extracted in November. Bands were separately excised and DNA from de agarose gel was extracted. The PCR product was used to construct libraries with a pGemTeasy cloning kit. The restriction fragment length polymorphism (RFLP) products were visualized after electrophoresis in low-melting-point agarose gel. Clones with different RFLP patterns were sequenced (Macrogen, Corea). Sequences of higher bands corresponded to eukaryotic organisms. However, sequences of lower DNA bands corresponded to archaeal organisms. Phylogenetic analysis showed that sequences of lower bands corresponded to eight different archaea. All of them were grouped with the Marine Group II of Euryarchaota organisms, which is characterized as a methanogen-halophile lineage. The organisms of this group have been shown to fix nitrogen and have been reported, in general, in the deep sea. This study relies in the importance of recovering rDNA archaeal sequences from surface water samples from the Argentinean Sea by using specific primers of eukaryotic organisms.