# Cyanobacteria as Chassis for Industrial Biotechnology: Progress and Prospects
## 3 Classical Genetic Engineering Technology  
### Cyanobacterial Transformation  
Transformation of cyanobacteria has been thoroughly reviewed [4; 85; 86; 22] and the methodology has been well described [5; 69]. The ability to transform cyanobacteria has been the drive for the development of the molecular biology of these organisms and has been the basis of many of their biotechnological applications [70]. Several cyanobacterial strains can be transformed with exogenously added DNA [4] or can accept mobilised plasmid DNA from _E. coli_ by conjugation [15]. Natural transformation was first described for _Synechococcus_ sp. strain PCC7942 [85] and hence is considered the primary means for gene transfer in unicellular cyanobacteria [6]. _Synechococcus_ and _Synechocystis_ species are among the few species that are known to be naturally transformable by exogenous DNA with little known about why natural transformation has thus far been restricted to certain unicellular cyanobacteria. This may be due to the presence of extracellular nucleases found in heterocyst-forming filamentous cyanobacteria that results in the degradation of the incoming DNA [71]. The natural transformable nature of _Synechococcus_ and _Synechocystis_ species explains why a lot of focus has been directed to the genetic engineering of these species. There are many factors that affect the frequency and efficiency of transformation in cyanobacteria, which can vary between \(10^{-3}\) and \(10^{-8}\)[4]. These include the used strain, the length, form and concentration of used DNA and also the method used for transformation [72]. The optimum conditions for transformation of the model organism _Synechocystis_ sp. PCC 6803 are well described [72; 73]. Since the discovery of cyanobacterial transformation in the 1970s, many techniques such as electroporation and ultrasonic treatment have been developed and applied to several cyanobacterial strains [70; 73; 74]. Electroporation has been used in the transformation of _Anabena_ sp., _Synechococcus_ sp., _Synechocystis_ sp., _Fremylla diplosphon_ and _Plectonema boryanum_, [70; 73; 74]. Ultrasonic transformation has been employed in the transformation of _Synechocystis_ sp. [70; 73; 74]. It was noted that, in _Synechocystis_ 6803, exogenous DNA is integrated into the genome following natural transformation by means of a well-established double homologous recombination system [72; 73]. Cyanobacteria that use systems with homologous recombination mechanisms allow for more effective gene modifications to wild-type genes [9]. This is true since gene replacement depends on homologous recombination in order to replace a section of the recipient chromosome with a donor DNA that contains a selectable marker [75]. [PERSON] and [PERSON] [76], were the first to demonstrate homologous recombination in cyanobacteria as a means of transformation, while [PERSON] and [PERSON], [77] were the first to achieve homologous recombination in _Anabaena_. Although the transformation of cyanobacteria was thoroughly investigated, very little is known on the mechanism by which DNA binds to the cell, gets processed and subsequently transported across the membrane during transformation [10; 72; 78]. However, it is thought that type IV pili are involved in the uptake process [79; 86]. Studying the effect of deleting the gene(s) encoding type IV pili may give an insight into their role in the transformation process. There is no doubt that the development of tools for the modification of cyanobacteria will help accelerate cyanobacterial research which will ultimately improve its use for industrial biotechnology applications. However, such improvements should not be restricted to the highly investigated unicellular model organisms but extended to include strains with desired characteristics such as those with fast growth rates, resistance to high temperatures and salinity, or resistance to infection and nitrogen fixers to mention a few.