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Method of ELISA applicable to toxoplasma gondii

A technology of Toxoplasma gondii and protein, which can be used in recombinant DNA technology, introduction of foreign genetic material using vectors, and material inspection products, etc., and can solve problems such as unclear key molecules.

Active Publication Date: 2018-05-18
SHENYANG AGRI UNIV
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  • Claims
  • Application Information

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Problems solved by technology

[0003] The key molecule of Toxoplasma gondii causing host pathogenicity is still unclear

Method used

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  • Method of ELISA applicable to toxoplasma gondii
  • Method of ELISA applicable to toxoplasma gondii
  • Method of ELISA applicable to toxoplasma gondii

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Experimental program
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Embodiment 1

[0057] Embodiment 1: the establishment of pig Toxoplasma gondii ELISA detection method

[0058] 1. Extraction of total RNA from Toxoplasma gondii ME49 strain

[0059] a. Take out the worm sediment from -80°C, add 1 mL of sterilized PBS to wash, and centrifuge for 10 min at 2500 rpm.

[0060] b. After centrifugation and washing, discard the supernatant PBS, add 1mL Trizol to the precipitate, and repeatedly pipette to ensure that the worms can be completely broken and lysed.

[0061] c. Add 200 μL of chloroform to 1 mL of Trizol, shake for 15 s with a shaker, and let stand on ice for 3 min. d. After standing still, place in a centrifuge at 4°C, centrifuge for 15 minutes at 12000×g, draw 400 μL of the supernatant and place it in a new sterile EP tube.

[0062] e. Add the same volume (400 μL) of isopropanol, invert and mix for 1 min, and let stand on ice for 10 min.

[0063] f. After standing still, centrifuge in a centrifuge at 4°C for 10 minutes at 12000g / min, discard the sup...

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Abstract

The invention provides a method of ELISA applicable to toxoplasma gondii. The method is characterized by comprising the following steps of using TGME49_223140 His label recombinant protein and GST label recombinant protein to detect pig serum; conducting double enzyme digestion on pEASY Simple Blunt plasmid containing a target fragment and using the same restriction enzyme to conduct double enzymedigestion on expression carriers pET-28a and pGEX4T-1; converting the recombinant expression carriers pET-28a and pGEX4T-1 into engineering bacteria E.coilBL21(DE3) and E.coil BL21 respectively to express and purify protein; using the purified TGME49_223140 His label recombinant protein and GST label recombinant protein as covering protein respectively and building an ELISA detection method.

Description

technical field [0001] The invention relates to the acquisition, expression and purification of the TGME49_223140 gene of toxoplasma gondii and the establishment of an ELISA method. Background technique [0002] Toxoplasmosis is a zoonotic parasitic disease caused by Toxoplasma gondii (Toxoplasma gondii), which has a worldwide distribution; it is seriously harmful to people and animals with immune insufficiency and immunosuppression. [0003] The key molecules of Toxoplasma gondii causing host pathogenicity are still unclear. [0004] Toxoplasmosis threatens human health and the development of animal husbandry. Contents of the invention [0005] Purpose of the present invention: to establish a sensitive and effective ELISA detection method for Toxoplasma gondii. [0006] The technical scheme provided by the invention is as follows: [0007] 1 nucleotide sequence [0008] (1) Acquire nucleic acid sequence from TOXODB (http: / / toxodb.org / toxo / ) database. [0009] (2) The...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/70C12N15/66G01N33/96G01N33/68
CPCC12N15/66C12N15/70G01N33/68G01N33/96
Inventor 陈启军李烨馨姜宁杨娜桑晓宇冯颖
Owner SHENYANG AGRI UNIV
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