This research was aimed at estimating the size of the genome for native strains, and detecting extrachromosomal elements named megaplasmids containing important genes encoding enzymes involved in solvent production. This work is related to 13 native bacterial strains from the Clostridium genus, selected from a strain-bank (consisting of 178 isolates from different Colombian soils), based on their greater ability to produce solvents as acetone, butanol, ethanol and 1,3 propanediol.

The genus Clostridium is a heterogeneous genus of anaerobic bacteria. Clostridia contain many species known by its biotechnological potential to produce high-added value solvents. Those process has been exploited from the First World War on an industrial scale in the so-called Weizmann process (acetone-butanol-etanol ABE) fermentation. However due to the foreseeable shortage of fossil energy sources, fuels from renewable sources are gaining increasing interest.

On the other hand, research aimed at gaining knowledge about genetic material has often relied on estimating genome size. PFGE is a molecular technique used for estimating the genome size of several bacteria. In order to determine the genome size, bacterial chromosome is cut with enzymes, and the fragments are then separated by PFGE. Finally the size of the fragments is determined and the obtained values are added for calculating the entire genome size.

Many efforts have been made in recent years to improve and standardise PFGE procedures for determining genome size in different bacteria. They generally consist of immobilizing cells in agarose plugs, breaking cell walls with enzymes to expose DNA, cutting it and separating fragments by electrophoresis. In this work three parameters were taken into account to standardise the method: establishing a DNA concentration to give good images of the fragments, controlling agarose concentration and using at least three hydrolytic treatments for obtaining DNA.

In our work mean genome size determined for the C. acetobutylicum ATCC 824 reference strain was 4,136 ± 169 Kpb, which is comparable to the size estimated for the same strain previously. Genome sizes for Colombian clostridia strains were ranged from 4.0 to 4.2 Mbp (Table 1). Only one extra-chromosomal element was detected in our work as migrating between 145.5 kb and 194 kb marker bands in all native strains.

A region responsible of solvent production (adc gene) was detected in one native strain by using PCR techniques. The adc gene encodes acetoacetate decarboxylase which plays a fundamental role in solventogenic Clostridia's metabolic route. This fact suggests that genes related to solventogenesis in native strain IBUN18A may be localised in the extra-chromosomal element detected in this work.

The PFGE method developed in this study was shown to be useful for obtaining estimated genome sizes for thirteen Clostridium strains isolated from Colombian soils. Further analysis is required for detecting solventogenic genes in native strains. Determining genome size and the physical structure of bacterial genomes could provide an alternative parameter to be employed in taxonomic studies. Results obtained in this work are useful for increasing genetic information about native Colombian strains.