The Primary Research Of All0012 In Anabaena PCC7120

Nowadays,cyanobacteria blooms have been occurring in lakes worldwide and have been reported to have harmful effects on human being. In freshwaters , the environment problem of cyanobacteria blooms are the result of an excess of nutrients we commonly call eutrophication. At present, while controlling of water pollution, many researcher try to find the way to prevent cyanobacteria excess growth. It become the main research direction in the field of water pollution control.The cpcM genes of Synechococcus sp. strain PCC 7002 and Synechocystis sp. strain PCC 6803 were insertionally inactivated,it encodes the enzyme that methylates the amide nitrogen of Asn71/72 in CpcB, ApcB, and ApcF.This N-methylation of Asn72 is important for efficient energy transfer in PBS.In this paper, by using a comparative bioinformatics approach, we found highly conserved open reading frames, all0012 in Anabaena sp. strain PCC 7120, which is homologous to cpcM and it also encode a previously uncharacterized methyltransferase family.In this work, we used molecular cloning methods to restruct a plasmid vecter pBlue-L-str/sp-R, which containing the homologous arms of all0012 gene fragment, and streptomycin resistance gene was insert in it with the appropriate restiction sites, then put the fragment contains the homologous arms and streptomycin resistance gene into a integrative plasmid vector pRL271. After that, we transformed the recombinanted vector into E.coli. The final construct was conjuncted into Anabaena sp. strain PCC 7120, the recombinant was selected by Streptomycin and Spectinomycin BG11 plate screening.We compared the deletion mutant of Anabaena sp. strain PCC 7120 with the wild type at different light intensity. At a low light intensity,there is no significant difference between mutant and wild type with their doubling times. The all0012 deletion mutant grows slowly at high light intensity and sensitive to the high light intensity, so we all0012 gene could code the enzyme that methylates the amide nitrogen of Asn71/72 or at least involved in this procession.In addition, synthesis of the desired protein in the recombinant E. coli system was tested and validated by SDS-PAGE. The results show that selecting the right gene, all0012 gene in vivo in the corresponding recombinant heterologous expression system to express the same molecular weight as expected and the expression product present in the supernatant.