Team:ECUST China/Proof Of Concept
Our project is theoretically feasible, but we have to admit that we have not achieved what we describe in Implementation. In Result, we have described the success of our experiment. Our previous verification experiments were conducted in two modules, DNA Walker and E-CRISPR, both of which have been able to achieve qualitative detection.
The purpose of DNA Walker is to amplify the input nucleic acid signal, and we used 30nt ssDNA as the target to verify the amplification effect of DNA Walker. The experimental results showed that when a certain amount of target was input, DNA Walker would feedback a higher amount of C:F double strand as the output signal, which was consistent with our expectation. Today we are still optimizing the experimental conditions to achieve the minimum signal input, output stability, and considerable C:F double chain.
The C:F double chain we design will encounter a modified G chain on the electrode after entering the E-CRISPR module, and the G chain will compete with the F chain for the C chain. We have achieved this competition in the EP tube, which shows that we can theoretically do this on the electrode as well.
The purpose of E-CRISPR is to achieve the output of electrical signals by cutting the G chain, which includes two factors: one is the cutting of G chain, and the other is the detection of signals. In the EP tube, we found that the input of C chain single strand can trigger the cutting of Cas9 Nickase, making the free G chain be cut off. At this point, the fragment of G chain can be detected by PAGE, and our electrophoresis results are good proofs of the occurrence of cutting. In the experiment on the electrode, after the G chain modified on the electrode plate is cut, we can see the decrease of the electrical signal, which indicates that it is feasible to detect the cutting of the G chain with the electrode.
In the above three steps, we have verified the theoretical feasibility of D-E-Tector through different modules. Now we need more time for the joint verification of the two modules to realize the real secondary amplification and detection. We will also combine the two modules through DMF devices to achieve simultaneous multi-channel detection. Finally, we hope to verify the actual detection capability of D-E-Tector by using Pseudovirus or even relying on hospitals to test the infected people.
In the above three steps, we have verified the theoretical feasibility of D-E-Tector through different modules. Now we need more time for the joint verification of the two modules to realize the real secondary amplification and detection. We will also combine the two modules through DMF devices to achieve simultaneous multi-channel detection. Finally, we hope to verify the actual detection capability of D-E-Tector by using Pseudovirus or even relying on hospitals to test the infected people.