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Babesia microti Bm186 antigen and application thereof

A technology of bm186 and Babesia, applied in the field of bioengineering, can solve the problem of low sensitivity of antigens and methods, and achieve the effect of preventing Babesia infection with high specificity and sensitivity

Active Publication Date: 2015-08-19
STATION OF VIRUS PREVENTION & CONTROL CHINA DISEASES PREVENTION & CONTROL CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The object of the present invention is to provide a kind of Babesia microti Bm186 antigen and application thereof, described this Babesia microti Bm186 antigen and application thereof have solved the antigen and method sensitivity of diagnosis Babesia microti disease in the prior art high technical issues

Method used

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  • Babesia microti Bm186 antigen and application thereof
  • Babesia microti Bm186 antigen and application thereof
  • Babesia microti Bm186 antigen and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Example 1 Collection of Babesia vole:

[0045] Babesia microti selected from the Babesia microti provided by the Institute of Experimental Zoology, Chinese Academy of Medical Sciences PRA-99 TM Insect strain. Babesia microti was injected intraperitoneally, and BALB / c mice were inoculated. About 4 days later, the infection rate of protozoaemia reached 20%. Anticoagulant whole blood was collected. Leukocytes were separated from erythrocytes using Percoll density gradient centrifugation to remove leukocytes. The erythrocytes were lysed by adding erythrocyte lysate, and the precipitate was collected after centrifugation, which was determined to be Babesia worms by microscopic examination.

Embodiment 2

[0046] The separation and purification of embodiment 2 mRNA:

[0047] The total RNA of Babesia microti was extracted with TRIzol reagent from Invitrogen Company, and the mRNA was isolated and purified from the total RNA according to the instructions of the Oligitex mRNASpin-Column Kit kit. And measure by agarose gel electrophoresis and ultraviolet analysis, detect the quality of mRNA (such as figure 1 shown).

[0048] Example 2 Transcriptome sequencing and analysis of Babesia microti:

[0049] The sequencing and analysis of the transcriptome were completed by Shanghai Ouyi Biomedical Technology Co., Ltd. The SMART cDNA library for sequencing was constructed using the SMART cDNA Library construction Kit kit from Clontech. Large-scale transcriptome sequencing was performed using a Roche 454 sequencer.

[0050] Preprocess the raw data of the transcriptome, remove the adapter sequence and primer sequence, and remove low-quality and short-length sequences. A total of 708,175 p...

Embodiment 3

[0052] Example 3 Search for candidate antigen genes:

[0053] For the 9377 Isotigs and 23088 Singlets in the transcriptome, first use the Tandem Repeats Finder software (version 3.3.0.0) to find tandem repeats, and set the parameters: "Alignment Parameters [match, mismatch, indel]" to 2, 7, 7 ; "Minimum Alignment Score To Report Repeat" is 50; "Maximum Period Size" is 2000. Then use NCBI's Open Reading Frame Finder (http: / / www.ncbi.nlm.nih.gov / projects / gorf / ) to find the cDNA sequence with a Minimum Alignment Score To Report Repeat value greater than 50; The amino acid sequence was predicted to contain signal peptide sequence using SignalP 4.1 software (http: / / www.cbs.dtu.dk / services / SignalP / ).

[0054] Using the above strategy, a total of 8 assembled sequences containing tandem repeat sequences and signal peptide sequence structures were found in the transcriptome of Babesia microti. Among them, the sequence of Isotig00186 was compared by BLAST, and it was a novel gene. The...

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Abstract

The invention provides a separated gene, the sequence of which is as shown in SEQ ID NO: 3. The invention further provides a babesia microti Bm186 antigen protein, which is encoded by the separated gene, and the amino acid sequence of the antigen protein is as shown in SEQ ID NO: 4. The invention further provides an antibody, which specifically combines the babesia microti Bm186 antigen protein. According to expression and purification of the babesia microti Bm186 antigen, a high specificity and sensitivity babesia microti diagnostic method is further established, and the antigen can be applied to different aspects, such as scientific research, and prevention and control of babesia microti; a vaccine provided by the invention has an effect of preventing babesia microti infection. A kit and a colloidal gold immunochromatographic test strip, which are provided by the invention, further can be used for detecting the babesia microti.

Description

technical field [0001] The invention belongs to the field of bioengineering, and relates to a parasite Babesia vole, specifically Babesia vole Bm186 antigen and application thereof. Background technique [0002] The first confirmed case of human babesiosis was reported in Yugoslavia in 1957. Subsequently, multiple cases of human babesiosis were reported successively in many European countries and the United States, and a small number of cases also appeared in Japan, mainland China and Taiwan in Asia, South Africa and Egypt in Africa. Babesia is a tick-borne zoonotic disease caused by Babesia parasitic on the red blood cells of vertebrates. At present, the Babesia that can cause human babesiosis have been identified, including a few species of Babesia (Babesiidae) such as Babesia microti and Babesia divergens. . [0003] Humans are usually infected through the bites of carrying ticks or clinical transfusion of blood donated by asymptomatic infected persons. The clinical s...

Claims

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Application Information

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IPC IPC(8): C12N15/30C07K14/44C07K16/20C12Q1/68A61K39/018A61K39/395A61P33/02G01N33/68
Inventor 许学年周岩程娜卢艳徐斌陈韶红胡薇
Owner STATION OF VIRUS PREVENTION & CONTROL CHINA DISEASES PREVENTION & CONTROL CENT
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