Human papilomavirus typing gene chip detecting system
A technology of human papillomavirus and gene chip, which is applied in the field of gene chip; can solve the problem that the limitation of detection flux cannot meet the clinical needs and other problems
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Embodiment 1
[0095] [Example 1] Preparation of HPV Typing Detection Chip
[0096] 1. Manufacturers and specifications of raw materials
[0097] name
manufacturers
Specification
Product name
EDAC
Sigma
Nylon membrane
Pall
Pore size: 0.45uM
30cm×3M
Biodyne C
Membranes
[0098] The probe sequence design is shown in SEQ ID 1-36, and the 3'-terminal amino-labeled probe was synthesized by Shanghai Boya Biotechnology Co., Ltd. according to methods well known to those skilled in the art.
[0099] 2. Production of membrane strips
[0100]
Embodiment 2
[0101] [Example 2] Sample detection of HPV typing chip detection kit
[0102] From August to October 2003, clinical verification was carried out in Shenzhen Hospital of Peking University and Shenzhen People's Hospital respectively. The results proved that the detection system of the present invention has large throughput and high sensitivity. Quantitative PCR comparable sensitivity. The specific reagents and detection methods are as follows:
[0103] ●Kit composition
[0104]
[0105] Wherein the PCR Mix contains primers of SEQ ID NO.37-38.
[0106] ●Necessary equipment and prepared reagents
[0107] PCR instrument DNA electrophoresis instrument
[0108] Molecular Hybridization Box Shaker
[0109] 3%H 2 o 2
[0110] Other required reagents were prepared according to the following formula:
[0111] 20×SSC (the pH value of the solution directly affects
sound color results)
NaCl 175.3g
Sodium citrate 88.2g
Add H 2 O dissolved to 1000mL, ...
Embodiment 3
[0144] [Example 3] Comparison of the hybridization effect of the 3'-terminal amino-labeled probe and the 5'-terminal amino-labeled probe
[0145] The hybridization probe can be amino-labeled at the 3’ or 5’ end of the oligonucleotide. The following experiments compare the hybridization effects of the 3’ and 5’ amino-labeled probes:
[0146] Synthesize 3'-terminal amino-labeled probes and 5'-terminal amino-labeled probes respectively, prepare nylon membranes with the method in Example 1, and carry out hybridization according to the method of Example 2 with positive control samples and negative control samples, the results are shown in image 3 ,Depend on image 3 The results show that under the same conditions, the labeling effect of the 3'-terminal amino group is better than that of the 5'-terminal labeling effect; specificity.
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