Design
Fig 1. Project flow chart of SNIPER E
In order to solve the difficulty of echinococcosis detection in hydatid disease epidemic areas, we designed this year's project-Specific New diagnostic Implement Pointing at Echinococcosis(SNIPER E), which means we will target the DNA of echinococcosis.
As mentioned earlier, hydatid disease areas are mostly in remote areas and the economic level is relatively backward.The tranditional PCR amplification technology requires specialized equipment and takes a long time.
To solve this problem, we consulted the literature and some professors and decided to adopt the Recombinase Polymerase Amplification (RPA) method, which doesn't need any equipment, and the reaction time is much shorter than PCR.
Fig 2. Electrophoresis plots of SE plasmid amplified by different primers
CRISPR-csa12a system is a relatively mature DNA detection system, which can detect the DNA fragments we want according to the designed sgRNA, and has been widely used in our laboratory. We imagine that cfDNA is extracted from the patient's blood and then amplified with RPA.
Fig 3. Cas12a detection using different sgRNA
The amplified DNA was detected by colloidal gold strip after passing the CRISPR-CAS12A system. If the patient is not infected with echinococcosis, only C-band will appear on the test strip; if the patient is infected with echinococcosis, both C-band and T-band will appear on the test strip.
Fig 4. Determining the result of the test strip
At present, our design has been verified in the laboratory, and we can successfully detect hydatid cfDNA in patient samples.
