Basic & Clinical Medicine ›› 2022, Vol. 42 ›› Issue (1): 139-144.

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Direct detection of Plasmodium female gametocyte-specific transcript s25 mRNA in blood by CLIP-PCR

  

  • Received:2021-10-08 Revised:2021-11-22 Online:2022-01-05 Published:2022-01-05

Abstract: Objective To establish a molecular detection method for the Plasmodium female gametocyte. Methods Capture and ligation probes was specific designed based on Plasmodium falciparum and Plasmodium vivax femal gametocyte special RNA targets,which is s25 mRNA. After the whole blood is lysed, the RNA target is directly released and fixed on the 96-well plate with capture probes without extraction. Washing away the unbound probe, the ligation probes which also hybridized to the RNA target is specific ligated to form an single-strand amplification template with specific-designed sequence at both ends. The SYBR Green qPCR was carried out with universal primers and the TaqMan qPCR was carried out with universal primers and universal fluorescent probes. Evaluate the sensitivity, specificity and repeatability of this method and compared with the RT-qPCR methods. And it was also used to detect clinical samples. Results This method was successfully applied to detect s25 mRNA with high sensitivity and specificity. All three methods can detect as low as 11 copies of S25 mRNA targets, and this method is simpler than RT-qPCR. And it can accurately detect gametocytes in the blood of malaria patients. The detection of 96 samples can be completed within 3h. Conclusion Established a method for the femal gametocyte of Plasmodium falciparum and Plasmodium vivax detection which provides a sensitive and efficient method for large-scale disease screening of gametocytes and control of malaria transmission .

Key words: Plasmodium, gametocyte, detection method

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