Difference between revisions of "Part:BBa K1217003"

Revision as of 06:06, 5 October 2013

pT7-ppk1(K.oralis)

Polyphosphate kinase (ppk1) (E.C. 2.7.4.1) catalyzes the formation of polyphosphate from ATP. In BBa_K1217003, ppk1 was under the control of T7 promotor. Under iptg induction, over-expression of ppk1 can reduce the phosphate level in the medium by its incorporation into polyphosphate synthesis inside the bacteria.

Usage and Biology

Efficiency of this construct is compared with Two other designs BBa_K1217008 and BBa_K1217010. We test the constructs' efficiency from 2 parameters: poly-P synthesis efficiency and phosphate removal efficiency from the medium.

• 

  Expected model of BBa_K1217003

• 

  Expected model of BBa_K1217008

• 

  Expected model of BBa_K1217010

Expression of BBa_K1217003

We confirm the expression of PPK1 enzyme in e.coli.

SDS-PAGE analysis of expression of K.oralis PPK1 enzyme inside E.coli. Expression of PPK1 is induced by 0.5mM IPTG at 30 degree for 5 hours. Band marked with star is the PPK1 enzyme, the enzyme expressed is soluble.

Efficiency of poly-P synthesis and phosphate removal from environment

Experiment procedure. Wide type and blank act as a control to indicate the normal phosphate uptake of wide type e.coli and the phosphate level change along with time. Upon iptg induction, cultural samples are collected in every three hours. The cell pellet harvested is used to measure the intracellular poly-P level and the supernatant is used for phosphate quantification

Intracellular poly-P quantification

Phosphate level in the medium Testing

Sequence and Features