The directed modification of DNA through recombinant DNA technology, widely known as genetic engineering, represents a powerful tool not only for improv­ing strains of industrial microorganisms (e. g., by the increase of product yields), but also for transferring specific traits or properties from some species to oth­ers, which are very separated in the evolutionary chain. This technology implies the extraction of genes from organisms exhibiting a determined trait in order to introduce (recombine) them in the DNA of a host microorganism (or organism). When the modified microorganism is reproduced, the succeeding generations will inherit the newly acquired trait. In the case of bacteria, the most common vectors used for DNA transfer to the host organism are the plasmids, relatively short autonomous (nonintegrated to the bacterial chromosome) DNA sequences that are autoreplicable, i. e., can form copies of themselves especially during the cell reproduction. It is worth emphasizing that the recombination of the gene with the plasmid is carried out in vitro (Figure 6.4). The recombinant vector obtained is introduced into the bacterial cells through diverse techniques that include a temporal increase in the porosity of both cell wall and cell membrane for the vec­tor to be introduced into the cytoplasm attaining, in this way, the transformation of the host cells. The microorganisms having genes of other organisms are called recombinant microorganisms or, in a general way, genetically modified organ­isms or simply engineered organisms.

Ffinrifir ritnr fnr DMA chain hyrlrnlyr

Foreign DNA chaii DNA after cleavage

Vectors replication

Cell reproduction

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FIGURE 6.4 Principle of recombinant DNA technology to develop genetically modified bacteria. A plasmid is the vector for transferring the foreign gene to the bacterium.

The introduction of recombinant microorganisms in the ethanol industry can lead to the development of industrial processes radically different from the conven­tional technologies based on the fermentation of molasses or starch hydrolyzates using native or improved yeast strains. Due to the specific and directed charac­ter of the modifications that can be done by genetic engineering, it is possible to “create” microbial strains capable of assimilating alternative feedstocks (first, lignocellulosic materials) or ones with the ability to perform simultaneous trans­formations in integrated process (e. g., the direct conversion of starch or lignocel — lulosic biomass). In this regard, when the microorganisms are genetically modified
to assimilate more substrates or to form new products, the integration of a series of very complex chemical transformations allowing the execution of several chemical processes in the same unit can be carried out. This kind of integration is verified at cell or even molecular level. These integrated processes represent a new approach in ethanol production and are the base for integral utilization of feedstocks in the so-called biorefineries (Cardona and Sanchez, 2007).