Construction of Local Isolates of Cyanobacteria for Ethanol Production.


Payam Bayazeed Hasan and Dlnya Asad Mohamad

Faculty of Science and Science Education- Sulaimani University- Qliasan- Sulaimani- Iraq



Abstract

Cyanobacteria can use solar energy and convert carbon dioxide into biofuel molecules in
one single biological system. In this research, Synechococcus sp. was isolated from
Saray Subhan Agha fresh water, a pure culture of Synechococcus was obtained by
several subculturing on BG11 media. For the production of ethanol by Synechococcus
sp. pyruvate decarboxylase (PDC) and alcohol dehydrogenseII (ADH II), genes from
Zymomonas mobilis ATCC (29191), were amplified by PCR and cloned into the
pSyn_1⁄D-TOPO® Vector. The Synechococcus and Synechococcus elongates
transformed with constructed vector (pSyn_1⁄D-TOPO®) that harboring the two ethanol
fermenting genes. The transformation was performed using a double homologous
recombination system to integrate the PDC and ADHII genes into the local isolates of
Synechococcus sp. and Synechococcus elongatus chromosome under the control
cyanobacterial weak constitutive nickel inducible promoter. The recombinant
Synechococcus cells grow in different concentrations of NiSO4 (1, 2.5, 5, 7.5, 10) μM in
BG11 media, under different temperature (15, 30, 45)  ̊C and different light intensity (10,
50, 150) μE. The enzymatic ethanol assay kit was used to determine ethanol
concentration produced by both recombinant Synechococcus sp. and recombinant
Synechococcus elongatus. Highest ethanol concentration obtained by those cultures
containing five μl NiSO4, which incubated under continues light of 50μE at 30 ̊C
(Optimum um condition for ethanol production by recombinant Synechococcus cells).
The amount of ethanol produced by local isolates of Synechococcus sp. was 0.00103 g/l,
whereas for Synechococcus elongatus was 0.0138 g/l. The amount of ethanol produced
by those Synechococcus cultures containing different concentrations of NiSO4 were
incubated under continuous light of (10 and 150) μE and temperature of (15 and 45)  ̊C
was less than those cultures were incubated under light of 50 μE and temperature of
30 ̊C.

Key Words:
cyanobacteria,
Synechococcus,
Ethanol, Sulaimani.


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