[Construction of an engineered Acidithiobacillus caldus with high-efficiency arsenic resistance].

Using the recombinant technique in vitro, a new arsenic resistance plasmid pSDRA4 was constructed by subcloning the arsenic resistance genes from plasmid pUM3 into the wide-host-range IncQ plasmid pMMB24 with the hybrid trp-lac ( tac ) promoter, and followed by deleting the regulative gene of the promoter, the lacIQ gene. Then plasmid pSDRA4 was introduced from E. coli into extremely acidophilic obligately chemolithotrophic Acidithiobacillus caldus by conjugative transfer with a frequency of( 1.444 +/- 0.797) x 10(-4), and the engineered strain of Acidithiobacillus caldus (pSDRA4) for biomining was constructed. The successful transfer demonstrates the development of a conjugational system between strains of E. coli and A. caldus. The recombinant plasmid pSDRA4 is stable in A. caldus. Compared with wild type A. caldus, the level of the arsenic resistance of A. caldus (pSDRA4) is greatly raised from 10mmol/L to 45mmol/L.

Conversion of an obligate autotrophic bacteria to heterotrophic growth: expression of a heterogeneous phosphofructokinase gene in the chemolithotroph Acidithiobacillus thiooxidans.

A plasmid pSDK-1 containing the Escherichia coli phosphofructokinase-1 gene (pfkA) was constructed, and transferred into Acidithiobacillus thiooxidans Tt-7 by conjugation. The pfkA gene from E. coli could be expressed in this obligately autotrophic bacterium but the enzyme activity (18 U g-1) was lower than that in E. coli (K12: 86 U g-1; DF1010 carrying plasmid pSDK-1: 97 U g-1). In the presence of glucose, the Tt-7 transconjugant consumed glucose leading to a better growth yield.