A detection method for the diagnosis of snail infection, an intermediate host of Schistosoma japonicum

A technology for infection detection and schistosomiasis, applied in biochemical equipment and methods, microbial measurement/testing, DNA/RNA fragments, etc., can solve problems such as lack, achieve high accuracy, good application prospects and promotion value, and reduce interference Effect

Active Publication Date: 2021-06-08
SUN YAT SEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Currently, there is a lack of an efficient method for the detection of Schistosoma japonicum infection in snails

Method used

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  • A detection method for the diagnosis of snail infection, an intermediate host of Schistosoma japonicum
  • A detection method for the diagnosis of snail infection, an intermediate host of Schistosoma japonicum
  • A detection method for the diagnosis of snail infection, an intermediate host of Schistosoma japonicum

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Example 1 Primer Design

[0047] 1. Experimental method

[0048] According to the 28S sequence of Schistosoma japonicum (GenBank: Z46504.4, the nucleotide sequence is shown in SEQ ID NO: 7), a specific primer combination suitable for LAMP was designed and screened for specificity, sensitivity and amplification efficiency.

[0049] 2. Experimental results

[0050] According to the screening of specificity, sensitivity and amplification efficiency, a group of specific primer combinations for detecting LAMP infected by Schistosoma japonicum was determined, and its sequence is as follows:

[0051] Outer primer pair F3 (SEQ ID NO: 1): 5'-AAGTCGGAAACCGCTAAGG-3';

[0052] Outer primer pair B3 (SEQ ID NO:2): 5'-GCTTACGCCTGAGGCTTC-3';

[0053] Inner primer pair BIP (SEQ ID NO:3):

[0054] 5'-GCTGGGCCTATGCTGTGTGACCTCCTACTCGTTGCCAT-3';

[0055] Inner primer pair FIP (SEQ ID NO:4):

[0056] 5'-GGTATAGGTCCAACGCTCGAGCGTGTAACAACTCACCTGCCG-3';

[0057] The loop-promoting primer ...

Embodiment 2

[0062] Example 2 A kit for detecting Schistosoma japonicum infection

[0063] 1. Composition

[0064] Nucleotide sequence such as the primer set shown in SEQ ID NO:1~6 and 10×Reaction buffer, MgSO 4 (100mM), dNTP Mix(10mM), ddH 2 O, Bst DNA polymerase. .

[0065] Second, the method of use (such as figure 2 shown)

[0066] 1. Extraction of DNA from samples to be tested:

[0067] 1) Collect samples to be tested;

[0068] 2) Add 200 μl of proteinase K lysate to the sample to be tested in step 1), and digest in a water bath at 55° C. for 4 to 8 hours;

[0069] 3) Take out the sample in step 2), add an equal volume of phenol: chloroform: isoamyl alcohol with a volume ratio of 25:24:1 for extraction, take the water phase and add an equal volume of isopropanol for precipitation;

[0070] 4) Wash the precipitate collected in step 3) with 100 μl of 70% ethanol, then centrifuge at 8000 rpm for 8 min, discard the supernatant, and dry the DNA sample naturally;

[0071] 5) Dissol...

Embodiment 3

[0084] Embodiment 3 detection sensitivity effect analysis

[0085] 1. Experimental method

[0086] The shell of the oncomelania sample was removed, and 30 mg to 50 mg of oncomelania meat was taken as the sample to be tested, and the sample DNA of different concentrations was detected by LAMP using the kit of Example 2, and the SjCaBP, ShDral, SjR2, and Sm1-7 genes were used as controls, and the amplified The primers are:

[0087] wxya

[0088] SJ CBP-F3 GACCTACCAGAAAACAGCT SJ CBP-B3 CCATTAAGACATTTTCGATCATGT SJ CBP-FIP TCACCTGTGTATATTTCATGTGCAT-AACTGGGTTGAAATAATCACCA SJ CBP-BIP TAACCTGTATTACTGCCCACTCA-CAAATGAATGATCTTGTCACAC

[0089] ShDral

[0090] SH Drai-F3 gat ctc acc tat cag acg SH DraI-B3 GTC ACC AAT AAT ATG AAAC SH DraI-FIP CCACCAACAATTTTAAATTTTATCAGACGAAACAAAGAAAAT SH DraI-BIP GGTCGTATCGTTGTGAAATTTT CACCAATAATATGAAACAAT

[0091] J2

[0092] SJ SiR2-F3 TGGTTGAAAATTGATGGTAGT SJ ...

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Abstract

The invention discloses a detection method for diagnosing the infection of Schistosoma japonicum intermediate host snails. The invention uses a primer combination for detecting Schistosoma japonicum infection with a nucleotide sequence as shown in SEQ ID NO: 1-6 to establish a Schistosoma japonicum infection testing methods and testing kits. The invention can quickly and accurately detect the schistosomiasis in the inspected snails, and is simultaneously applicable to the detection of the infection of the schistosomiasis in animals and the monitoring of the schistosomiasis in the environment. The detection method has high detection rate, high sensitivity, strong specificity, high accuracy and simple equipment, so it has good application prospect and promotion value.

Description

technical field [0001] The invention relates to the technical field of parasite molecular detection, and more specifically relates to a detection method for diagnosis of oncomelania infection, an intermediate host of Schistosoma japonicum. Background technique [0002] Schistosomiasis is a zoonotic parasitic disease caused by schistosome infection that seriously affects human health and social and economic development. The schistosomiasis parasitic on the human body mainly include Schistosoma japonicum, S. S. haematobium), S. intercalatum, S. mekongi and S. malayensis. According to the statistics of the World Health Organization (WHO), about 200 million people in the endemic areas of 74 countries are still affected and threatened by schistosomiasis. According to statistics, as of the end of 2016, there were still about 54,000 schistosomiasis patients in my country, and the prevention and control situation in some areas was still grim. In my country, the main pathogen causi...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6888C12N15/11C12Q1/6844
CPCC12Q1/6844C12Q1/6888C12Q2531/119C12Q2563/107
Inventor 高江梅王立富曾幸达吴忠道
Owner SUN YAT SEN UNIV
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