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Rapid amplification method of hepatitis B virus nucleic acid

A hepatitis B virus and nucleic acid technology, applied in the field of biochemistry, can solve the problems of unfavorable acquisition of hepatitis B virus nucleic acid samples, complicated PCR methods, and restrictions on technology development, and achieve the goals of improving heat transfer efficiency, rapid and simple amplification, and excellent amplification efficiency Effect

Active Publication Date: 2018-11-30
SANSURE BIOTECH INC
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Problems solved by technology

As the basic technology of molecular biology research, PCR has promoted the development of life sciences, but the general PCR method is complicated and time-consuming, which limits the further development of the technology and is not conducive to the rapid acquisition of a large number of HBV nucleic acid samples

Method used

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  • Rapid amplification method of hepatitis B virus nucleic acid
  • Rapid amplification method of hepatitis B virus nucleic acid
  • Rapid amplification method of hepatitis B virus nucleic acid

Examples

Experimental program
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Effect test

Embodiment 1～16

[0048] According to the operation steps, 16 samples containing hepatitis B virus were amplified. The PCR reaction conditions were: pre-denaturation at 94°C for 1min; denaturation at 94°C for 0s, annealing and extension at 57°C for 0s, 40 cycles and heating at 57°C in each cycle Fluorescence collection was performed during the process to 94°C, and the total time of the amplification program was 15 minutes. The PCR machine used is the GNM-C7-8 real-time fluorescent quantitative PCR machine produced by Ginnomi Biotechnology Co., Ltd. Amplification curve such as Figure 5 As shown, it can be seen that the amplification curves of Examples 1-16 maintain a good shape and have high amplification efficiency, and the logarithmic values ​​(LOG values) are shown in Table 1.

[0049] In addition, the blank samples (samples not containing hepatitis B virus) were amplified according to the methods of Examples 1-16, and no false positive results occurred.

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Abstract

The invention relates to a rapid amplification method of hepatitis B virus nucleic acid. The method comprises the following steps: mixing a sample containing hepatitis B virus with a nucleic acid release agent, and then adding polymerase chain reaction (PCR) premix to obtain reaction liquid, wherein the nucleic acid release agent is prepared from surfactin, potassium chloride, sodium dodecyl sulfate and ethanol, and the PCR premix is prepared from deoxyribonucleoside triphosphate, an upstream primer shown in a sequence of SEQ ID No. 1, a downstream primer shown in a sequence of SEQ ID No. 2, DNA polymerase and an amplification buffer solution; putting the reaction liquid into a PCR reaction tube to form a thin film having thickness of 0.1 mm or less; putting the PCR reaction tube into a PCR amplification instrument and carrying out PCR amplification according to the following reaction conditions: carrying out pre-degeneration for 10-600s at 90-100 DEG C, carrying out degeneration for 0-1s at 90-100 DEG C, and annealing for 0-1s at 50-65 DEG C. Under the premise of guranteeing the accuracy and effectiveness of amplification, the rapid application method significantly shortens the time required for each cycle, thus achieving the aim of rapidly and simply amplifying the hepatitis B virus nucleic acid.

Description

technical field [0001] The invention relates to the field of biochemistry, in particular to a rapid amplification method of hepatitis B virus nucleic acid. Background technique [0002] Hepatitis B virus (HBV for short) is a DNA virus belonging to the family Hepadnaviridae. About 257 million people are infected with HBV in the world, and there are 93 million people infected with HBV in China. Therefore, research on hepatitis B virus has been paid attention to worldwide. In the related scientific research of hepatitis B virus, it is often necessary to amplify hepatitis B virus nucleic acid for non-disease diagnosis or therapeutic purposes, so as to obtain a large amount of hepatitis B virus nucleic acid, thereby providing samples for various scientific experiments. [0003] Polymerase chain reaction (PCR) is a major nucleic acid amplification technology in vitro, which has developed rapidly in recent years. The feature of PCR technology is to simulate the DNA replication p...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/686C12R1/93
CPCC12Q1/686C12Q1/706C12Q2563/107C12Q2527/113Y02A50/30C12Q2527/125
Inventor 范旭戴立忠
Owner SANSURE BIOTECH INC
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