Difference between revisions of "Team:NU Kazakhstan/Results"
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Revision as of 20:35, 17 October 2018
For the vector construction, SQR gene was cloned into pSyn_6 vector, and through gel electrophoresis was checked the gene assembly. Figure 1 illustrates the bands of cloned pSyn_6 (5577 bp) in 1-4 wells between 3 and 4 ladder bands, which confirms the success of gene assembly. Also, cloned pSyn_6 plasmid was tested on a presence of SQR gene by PCR amplification using SQR primers. In Figure 2, we can see SQR bands (1271 bp) between 8 and 9 ladder bands that evidences of SQR presence in cloned pSyn_6 plasmid.
Figure 1. Agarose gel electrophoresis (1%) of cloned pSyn_6 plasmid with SQR (5577 bp).
Figure 2. Agarose gel electrophoresis (1%) of PCR amplified products using SQR primers to test its presence in cloned pSyn_6.
After the gene construction, all attention was focused on transformation of cyanobacteria with SQR gene
Figure 3. Lamp with high light intensity.
Figure 4. Cyanobacteria colonies in BG-11 agar plates with spectinomycin.
Confirmation of integration of the cloned pSyn_6 plasmid into genome was done by colony PCR amplification using SQR primers. In Figure 5, we can see colonies with inserted SQR genes, which were used to get liquid genetically modified cyanobacteria culture.
Figure 5. Agarose gel electrophoresis (1%) of colony PCR amplified products using SQR primers.
We inserted Sulfide Quinone Reductase into the pSyn_6 сyanobacterial protein expression vector, which performs the function of converting toxic hydrogen sulfide into elemental sulfur. Firstly, we tested the survivability of our cyanobacteria in different concentrations of Na2S and oil. Secondly, to check the functionality of the SQR gene in cyanobacteria we were conducted Na2S reduction assay.
Survival test of cyanobacteria in the oilAs our main goal of the project is the bioremediation of oil wastewater, we tested the survivability of genetically modified and wild-type strain of cyanobacteria in different concentrations of oil in the solution. After 3 days of the experiment (Figure 6, 7 and 8), it was analyzed that cyanobacteria with SQR gene are doing very good in 0.1%, 0.5% and 1% oil solution compared to wild-type strain.
Figure 6. Survival test of cyanobacteria in 0.1% oil.
Figure 7. Survival test of cyanobacteria in 0.5% oil.
Figure 8. Survival test of cyanobacteria in 1% oil.
Figure 9. Survival test of cyanobacteria in different concentrations of Na2S.
Na2S reduction assay
Na2S used in the experiments was not pure. It came from industrial waste, which was taken from the MKA engineering company. According to them, Na2S content was no less than 60% and the rest were impurities including heavy metals. The stock solutions were prepared considering the weight percent of 60%.