Specific human gene fragment, primer and probe for detecting specific human gene fragment and application of specific human gene fragment

Abstract

The invention discloses a specific human gene fragment, a primer and a probe for detecting the specific human gene fragment and application of the specific human gene fragment, and particularly discloses a method comprising the following steps: screening the human specific gene fragment, designing, synthesizing and verifying the specific primer probe, constructing a standard plasma and acquiring the accurate relative molecular mass, drawing a standard curve and finally conducting qPCR (Polymerase Chain Reaction) quantitative detection. Therefore, the most important characteristic of the specific human gene fragment lies in that due to the quite strong specificity of the probe and the primer, a human cell in a sample mixed with other animal sources can be qualitatively and quantitatively detected; a carrier T5 is also known for the first time to be used in the construction of the standard plasma, the time is saved, and the cost is also lowered; and the various steps of a method are optimized, so that the detection is more accurate and sensitive.

Description

technical field [0001] The invention belongs to the technical field of biological detection of pharmacokinetics, in particular to a specific human gene fragment and its primer, probe and application. Background technique [0002] Since its official birth in 1985, PCR technology has been widely recognized by the life science community. After decades of continuous improvement, the PCR method has developed from a qualitative analysis method to a quantitative detection method. From the original gene that can only amplify a few kb to the present, it can amplify DNA fragments up to dozens of kb. So far, there have been more than a dozen PCR techniques, which have become the fundamental cornerstone of genetics and molecular biology. Among the many technologies, the real-time fluorescent quantitative PCR (qPCR) technology, which came out in 1996, is the most widely used. It is the most advanced nucleic acid molecular diagnostic technology for clinical use in the world today, and is...

AI Technical Summary

Problems solved by technology

The existing detection system mainly has the following defects: 1. The standard plasmid construction process is cumbersome: generally, intermediate plasmids are required for amplification, or enzyme digestion and recovery are required after cloning to connect the target vector; 2. The standard plasmid construction efficiency is not high enough: The standard plasmid constructed by linking the target sequence with the target vector often has high non-specific ligation products; 3. The calculation of the molar molecular weight of the standard plasmid is not accurate enough: usually the average of the four bases of ATCG is multiplied by the base number of the constructed plasmid The molecular weight is estimated; 4. The drawing of the standard curve is not accurate enough: the dilution factor of the standard plasmid is not enough, and the standard products used to draw the standard curve are too few (generally only about 5); copy, and the copy numbers in different parts are slightly different, which cannot correspond to the cell volume well; at the same time, because the ALU multi-copy sequence is short, it is not suitable to design corresponding primer probes, and it is difficult to use qPCR for absolute quantification

Images