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Biochip for streptococcus pneumoniae serotype detection, and its detecting method and kit

A detection technology for Streptococcus pneumoniae and serotype, which is applied in the field of gene chip for detection of Streptococcus pneumoniae serotype and its detection method and detection kit, can solve the problem of low similarity, achieve simple operation, strong repeatability, The effect of high accuracy

Inactive Publication Date: 2005-12-21
TIANJIN BIOCHIP TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Because the oligosaccharide unit processing enzyme has specific donors, acceptors and linkages, the similarity between the gene sequences encoding the oligosaccharide unit processing enzyme is very low

Method used

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  • Biochip for streptococcus pneumoniae serotype detection, and its detecting method and kit
  • Biochip for streptococcus pneumoniae serotype detection, and its detecting method and kit
  • Biochip for streptococcus pneumoniae serotype detection, and its detecting method and kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Example 1: Design and preparation of probes

[0044] 1. Sequence acquisition: The 16s rDNA sequence of Streptococcus pneumoniae and the wzy (type 3 is Cap3A) sequence of different serotypes downloaded from the Sanger Institute and GenBank public databases were sorted out with Artemis software.

[0045] 2. Probe design: Import the above sequence into OligoArray2.0 software, and set the parameters as follows: -n 20; -130; -L 40; -D 3000; -t 79; -T 90; -s 65°C; x 65 °C; -N 2 ; -p 33, -P 65; -m GGGGG CCCCC TTTTT AAAAA; -g 15. Run the program to design probes online.

[0046] 3. Probe screening: Select a probe with a length of 35bp±2bp and a Tm of 75°C±2°C from the output results, and use the method provided in Example 4 below to perform probe screening by hybridization experiments, and finally obtain the Specific and sensitive probes required for the invention of gene chips.

[0047] In a preferred embodiment of the present invention, 260 probes with a length of 35bp±2bp...

Embodiment 2

[0050] Example 2: Design and preparation of primers

[0051] 1. Sequence acquisition: Artemis software was used to sort out the 16s rDNA sequence of Streptococcus pneumoniae downloaded from the public databases of Sanger Institute and GenBank and the wzy sequences of different serotypes of Streptococcus pneumoniae (type 3 is Cap3A).

[0052] 2. Design primers: Import the above sequences into the primer design software Primer Premier 5.0 software, and set the corresponding parameters as follows: Search For: PCR Primers, Search types: Both.SearchRanges: Sense Primer 1 to 672, Anti-sense Primer 1 to 672 , PCR ProductSize: 100bp to 1000bp.Primer Length: 20bp±2bp.Search Mode: Automatic.

[0053] 3. Primer selection: Select primers with a Tm value of 50°C±5°C, a length of 20bp±2bp, Hairpin: NONE, Dimer: NONE, False Priming: NONE, Cross Dimer: NONE, and primers including the probe sequence from the output results. Individual probes were manually adjusted, and primers were appropriat...

Embodiment 3

[0057] Example 3: Gene Chip Preparation - Chip Spotting

[0058] 1. Dissolving the probes: the probes synthesized in Example 1 were respectively dissolved in 50% dimethyl sulfoxide (DMSO) solution, so that the final concentration of the probes reached 1 μg / μl.

[0059] 2. Adding plate: Add the dissolved probe to the corresponding position of the 384-well plate, 10 μl per well.

[0060] 3. Spotting: as figure 1The shown 57.5mm × 25.5mm × 1mm (length × width × height) clean aldehyde slide (CEL Associates, Inc.) was placed on the stage of the chip spotter (Spotarray 72), using SpotArray Control software (Tele chem smp3 stealty pin), run the program, press figure 2 The arrangement shown is spotted on the aldehyde-formed glass slide in the spotting area of ​​4.5 mm × 4.5 mm to form a medium-low density DNA micro-array, and the array arrangement rules in the four spotting areas on the glass slide are the same.

[0061] 4. Drying: Dry the spotted chips overnight at room temperatu...

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Abstract

The invention relates to one kind of gene chip used in the pneumonia streptococcus blood serum examination and its examination method and the reagent box used thereof. The gene chip includes the solid phase carrier and the oligonucleotide probe fixed on this carrier. The probe includes the DNA fragment selected from the streptococcus pneumoniae 16s rDNA and the DNA fragment selected from the polyase gene of streptococcus pneumoniae. Use designed primer to enlarge and mark the gene group of the sample to be detected, then to hybrid the gene group using the biological chip, the different type of serum of the streptococcus pneumoniae can be detected.

Description

technical field [0001] The invention relates to a gene chip, a detection method thereof and a test kit for detection, in particular to a gene chip for detecting the serotype of Streptococcus pneumoniae, a detection method and a test kit for detection. Background technique [0002] Streptococcus pneumoniae (Streptococcus pneumoniae) is the main pathogenic bacteria that causes high morbidity and high mortality among people of all ages in developing countries and developed countries. Sinusitis, meningitis, bacteremia and other diseases pose a serious threat to human health. Every year in the United States, about 500,000 people suffer from pneumonia caused by Streptococcus pneumoniae, about 60,000 people have otitis media, about 50,000 people have bacteremia, and about 50,000 to 6,000 people have meningitis. Pneumonia is also one of the common diseases in my country. Every year, 2.5 million people suffer from pneumococcal pneumonia and cause 125,000 deaths among them (mainly mi...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 王磊王荃王敏冯露
Owner TIANJIN BIOCHIP TECH CO LTD
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