Primer, probe and kit for detecting type-16 HPV (human papillomavirus)
A technology for HPV virus and detection primers, which is applied in the direction of microbial determination/inspection, microorganisms, and microorganism-based methods, and can solve problems such as the inability to detect multiple types of HPV viruses at the same time.
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Embodiment 1
[0163] Example 1 Construction of HPV quality control
[0164] (1) The 16 types of HPV fragments were ligated to prepare a pseudovirus.
[0165] The 16 types of HPV viruses were respectively amplified with corresponding specific primers (for example, HPV6 type virus template, using the HPV6 type upstream primer HPV6-F and downstream primer HPV6-R designed by the present invention to amplify, collect Product), the amplified products of 16 types of HPV virus were confirmed to be correct by gene sequencing and then packaged into pseudoviruses. The pseudoviruses were prepared and provided by Beijing Boheng Kechuang Biotechnology Co., Ltd.
[0166] (3) Dilute the HPV pseudovirus according to the concentration provided by Beijing Boheng Kechuang Biotechnology Co., Ltd., and dilute it to 1×10 5 copies / mL and 5×10 3 copy / mL, and then detected by ABI7500 fluorescent quantitative PCR instrument.
[0167] The result is as figure 1 Shown: the left is 1×10 detected by 16 kinds of HPV...
Embodiment 2
[0169] Example 2 The composition and use of the detection kit
[0170] 1. Kit composition
[0171] Reagent Element Dosage HPV lysate Anhydrous sodium carbonate, Tris-HCl, EDTA 50μL PCR reaction solution DNA polymerase, dNTPs, Mg 2+ 14.5μL negative control sterile saline
positive control Pseudovirus (1×10 5 copies / mL)
Weak Positive Control Pseudovirus (1×10 3 copies / mL)
Group A Primer Probe Mixture HPV6-F, HPV6-R, HPV6-P; HPV11-F, HPV11-R, HPV11-P 5.5μL Group B Primer Probe Mixture HPV16-F, HPV16-R, HPV16-P; HPV18-F, HPV18-R, HPV18-P 5.5μL Group C Primer Probe Mixture HPV31-F, HPV31-R, HPV31-P; HPV52-F, HPV52-R, HPV52-P 5.5μL Group D Primer Probe Mixture HPV33-F, HPV33-R, HPV33-P; HPV45-F, HPV45-R, HPV45-P 5.5μL Group E Primer Probe Mixture HPV35-F, HPV35-R, HPV35-P; HPV51-F, HPV51-R, HPV51-P 5.5μL Group F Primer Probe Mixture HPV39-F, HPV39-R, HPV39-P; HPV5...
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