Team:JMU Wuerzburg/Results

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Bioinformatics

To detect Plasmodium in general we aligned 2080 sequences from NCBI. Our primers and the probe has a consensus of more than 98% and in one case 94,9%. The oligos are located in the mitochondria in a regulatory region.
In the case of Plasmodium falciparum, we aligned 1012 sequences of Plasmodium falciparum and our primer set and the probe has a higher consensus than 97,5%. The location of the oligos is in a regulatory region of the mitochondria.

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Wet Lab

In general, we can detect Plasmodium in all five cultured Plasmodium strains of Plasmodium falciparum (FCR3, Mali2K, HB3, Dd2 and 3D7 from Tuebingen and Heidelberg). In addition, we can detect our Biobrick (BBa_K2614000) that includes a synthetic template as a positive control for the qPCR that detects Plasmodium in general (figure 1). We can detect less than 1000 copies of our synthetic template with a probe (FAM) and less than 50 copies with SybrGreen.

Figure 1: qPCR with cultured Plasmodium stems and our BioBrick as a control for Plasmodium general

Moreover, we are able to detect P. falciparum in all five cultured Plasmodium strains. Our BioBrick is a negative control for our qPCR that detects P. falciparum. With the probe (HEX) we detect less than ten copies of our synthetic template. (figure 2)

Figure 2: qPCR with cultured Plasmodium stems and our BioBrick as a control for P. falciparum.

In a multiplex system the two probes (FAM and HEX) also detect the five cultured Plasmodium strains. They detect them with nearly the same cq-mean which means that both qPCRs can detect Plasmodium. With this result we showed that multiplexing is also possible with the two primer/probe pairs (figure 3).

Figure 3: Multiplex qPCR with cultured Plasmodium stems and our BioBrick as a control for Plasmodium general and specific for P. falciparum

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Human practices

We organised different workshops from scientists about malaria or primer/probe design which were open for all interested students. There was also a pipetting test where more experienced students could give their knowledge to pipetting beginners.

At our “cake and science” events we presented our project to the public.

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Interlab

We participated successfully at the fifth International InterLaboratory Measurement Study in synthetic biology. At the iGEM 2018 Vibrigens InterLab Study from Marburg we also take place.

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Collaboration

We were able to establish a very pleasant and close collaboration with team Munich. They have mentored us with great care and reliability and answered all our questions. So, it was much easier to start a new iGEM team in Wuerzburg. We also received advice from different members of team Tuebingen who were so kind to exchange some valuable information and experience with us.