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Plasmodium gene diagnosis primer

A gene diagnosis, Plasmodium technology, primer set applied in the diagnosis of Plasmodium, Plasmodium vivax and Plasmodium ovale infection, field of Plasmodium falciparum

Active Publication Date: 2019-10-15
KUNMING MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] At present, in the domestic market, apart from immunological diagnostic methods, there are no clinical rapid diagnostic kits with higher sensitivity and convenient application at the grassroots level

Method used

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Sequences of related specific and conserved genes were retrieved from GeneBank for comparison, and conserved gene fragments with a length of 200bp to 500bp were selected, and multiple sets of primers were designed using LAMP Designer 1.15 (PREMIER Biosoft) software, which were screened out through manual comparison and experiments. The most efficient and sensitive primer set. The sequence-specific primer set comprises the oligonucleotide set containing the nucleotide sequence represented by SEQ 1 to 6, and is used to amplify the specific region of the Plasmodium mtDNA gene sequence; the oligonucleotide containing the nucleotide sequence represented by SEQ 7 to 12 Group, for amplifying the specific region of Plasmodium falciparum ActinI gene sequence; Containing the oligonucleotide group of the nucleotide sequence represented by SEQ 13 to 18, for amplifying the specific region of Plasmodium vivax 18S rRNA gene sequence; Containing SEQ The oligonucleotide groups of the nu...

Embodiment 2

[0028] Establishment of LAMP reaction system

[0029] 1. Materials and methods:

[0030]Between 2016 and 2018, blood samples were collected from 466 suspected cases of malaria. Standard strain 3D7, DD2, P. knowlesi Strain H strain DNA (provided by MR4 / ATCC (Manassas, VA)). The DNA of Toxoplasma gondii and Schistosoma japonicum was provided by the Department of Parasites, Kunming Medical University.

[0031] 2. DNA template preparation

[0032] Using the Roche High Pure PCR Template Preparation Kit, the insect blood was taken for DNA template preparation.

[0033] Nested PCR reaction: primers and reaction conditions refer to published literature (Sun Lingcong, Zhang Huaxun, Pei Sujian, Xia Jing, Wu Dongni, Lin Wen: Etiological diagnosis and analysis of the first imported Plasmodium ovale subspecies wallikeri in Hubei Province. International Journal of Laboratory Medicine 2016,37:1956-1958.)

[0034] 3. LAMP reaction

[0035] The primer set used comprises a primer set comp...

Embodiment 3

[0043] specificity test

[0044] Tests were performed on other parasite species and on different Plasmodium species to analyze their specificity. Use the primer set of SEQ 1 to 6; the primer set of SEQ 7 to 12; the primer set of SEQ 13 to 18; the primer set of SEQ 19 to 24 to carry out LAMP reaction on Schistosoma japonicum and Toxoplasma gondii DNA respectively, and the result is negative. Use the primer set of SEQ1 to 6; The primer set of SEQ 7 to 12; The primer set of SEQ 13 to 18; The primer set of SEQ 19 to 24 is respectively to five kinds of malaria parasite Plasmodium falciparum (Pf), Plasmodium vivax (Pv ), Plasmodium ovale includes two subspecies of Plasmodium ovale curtisi (Poc) and Plasmodium ovale wallikeri (Pow), Plasmodium malariae (Pm), and the DNA of Plasmodium knowlesi (Pk) carry out LAMP reaction, the result is as follows:

[0045]

[0046] Plasmodium mtDNA gene

[0047] The primer set comprising SEQ 1 to 6 cannot amplify Toxoplasma gondii and Schistosom...

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Abstract

The present invention discloses a plasmodium gene diagnosis primer. The primer a loop-mediated isothermal amplification (LAMP) primer for detecting / identifying whether specific plasmodium and three species of plasmodium are present in samples. A use of the primer set can simultaneously or separately detect or identify infections of plasmodium and / or one or more of plasmodium falciparum, plasmodiumvivax and plasmodium ovale, gains precious time for diagnosis and treatment of malaria in China, and provides helps for preventing, controlling and eliminating prevalence of the malaria.

Description

technical field [0001] The invention relates to the field of biological detection and identification, in particular to a primer set capable of rapidly and accurately detecting and identifying Plasmodium infection, including a primer set for diagnosing Plasmodium, Plasmodium falciparum, Plasmodium vivax and Plasmodium ovale infections. Background technique [0002] Rapid and accurate malaria diagnosis is a daunting challenge in malaria-endemic countries. The clinical manifestations of malaria caused by different Plasmodium are different, and the treatment options are different. Therefore, the diagnosis of malaria requires accurate diagnosis of the species. [0003] In my country, many imported malaria infections involve two or more species, and these mixed infections are often overlooked or underestimated. Failure to detect co-infections leads to inadequate malaria treatment and can lead to exacerbations of the disease and serious complications. Therefore, there is an urgen...

Claims

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Application Information

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IPC IPC(8): C12Q1/6893C12Q1/6844C12Q1/04C12N15/11
CPCC12Q1/6893C12Q1/6844C12Q2531/119Y02A50/30
Inventor 杨照青陈熙张家祺徐士玲耿劲婷黄亚铭黄超曾炜林向征杨照坤
Owner KUNMING MEDICAL UNIVERSITY
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