Huaman plasma DNA quantitative analyser

Abstract

The present invention relates to a quantitative detection analysis method of human plasma DNA level. Said method includes the following steps: selecting artificially-synthetic exogenous DNA series whose extraction efficiency is identical to or similar to that of human plasma DNA, but they are non-homologous, selecting housekeeping gene beta actin as determination gene, inserting artificially-synthetic exogenous DNA into carrier pMD18T and placing it in the bacteria to make culture, making blue and white spot screening, colony PCR identification, enzyme-cutting into linearity, placing the above-mentioned material into plasma, adopting double fluorescent quantitative gene amplification, fluorescent groups are respectively FAM and HEX, extracting known exogenous positive plasmid recovery efficiency, using computation formula to make computation so as to obtain the quantity of plasma DNA.

Description

technical field [0001] The invention relates to a DNA measurement method, in particular to a quantitative detection and analysis method for plasma DNA level. Background technique [0002] Human plasma DNA refers to free extracellular DNA (cell free DNA), also known as circulating DNA, which is always at a relatively constant level and maintains a balance. Only when the human body suffers from various diseases, such as: body inflammation caused by infection, large area tissue damage caused by trauma, failure of tissues and organs, autoimmune diseases and tumors, etc. due to a large number of cell death, nucleic acid is released from the cells This balance will be broken only when a large amount of internal organs enter the blood of the peripheral circulation, resulting in a substantial increase in human plasma DNA levels and abnormal DNA quality. [0003] Therefore, the quantitative detection and analysis of plasma DNA can be used for the diagnosis and severity assessment of...

AI Technical Summary

Problems solved by technology

Although some people have used real-time fluorescence quantitative detection analysis for a housekeeping gene in the human genome before to try to quantify the level of human plasma DNA, it is always difficult to do so; the reason is that the beginning of the detection process must be used The template (plasma DNA template) for quantitative detection of plasma DNA is extracted and enriched, which will inevitably lead to differences in the extraction recovery rate of different samples (in the same experimental batch and each sample in different batches), and the highly sensitive method used subsequently There are certain differences in the gene amplification efficiency of each sample by sex gene amplification technology, which can cause serious deviations in the detection results. Therefore, the real-time fluorescence quantitative detection technology for plasma DNA alone cannot accurately quantify the level of human plasma DNA. As a result, the real-time fluorescent quantitative gene amplification (real-time fluorescent quantitative PCR) detection technology of human plasma DNA has long been difficult to be used in the diagnosis of clinical diseases, the observation of treatment effects and the analysis of disease prognosis

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