CLONING AND EXPRESSION OF POLYPHOSPHATE KINASE GENE FROM PSEUDOMONAS PUTIDA GM6
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    Abstract:

    A strain of high-efficiency phosphorus accumulating bacteria,identified as Pseudomonas putida GM6,was used as research object,from which polyphosphate kinase gene was cloned and its roles in the phosphate transport system verified.A 528-bp fragment of ppk gene was successfully amplified firstly from GM6 genomic DNA using self designed primers corresponding to the well-conserved regions of reported ppk gene sequences.Then,its upstream and downstream sequences were cloned with the technique of self-formed adaptor PCR (SEFA-PCR).Three amplified sequences were put together and analyzed using the OMIGA program (version 2.0),and operon of the complete ppk gene ca.2220 bp was obtained (it has been deposited in the GenBank database under accession number DQ133537).The constructed recombinant expression strain of ppk gene E.coli BL21 (DE3) /pET29a-ppk was induced with IPTGfor three hours,and expression product,81 kDa in molecular weight,was observed.The strain removed 80% of the phosphorus in the solution while the CK strain only 18% in 12 h.Its P removing capacity,more than 40%,the sofar reported highest rate,indicates excessive expression of ppk gene in E.coli,which would lead to accumulation of a great deal of poly-P in E.coli in vivo,and removal of a large amount of phosphate from the medium.

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Guan Libo, Cai Tianming, Li Bo, He Jian, Li Shunpeng, Cui Zhongli. CLONING AND EXPRESSION OF POLYPHOSPHATE KINASE GENE FROM PSEUDOMONAS PUTIDA GM6[J]. Acta Pedologica Sinica,2007,44(4):727-733.

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History
  • Received:March 31,2006
  • Revised:July 17,2006
  • Adopted:
  • Online: February 25,2013
  • Published: