Primer, kit and method for detecting staphylococcus aureus through PCR(Polymerase Chain Reaction)-pyrophosphate method

Abstract

The invention discloses a primer, kit and method for detecting staphylococcus aureus through a PCR (Polymerase Chain Reaction)-pyrophosphate method. In the invention, a PCR primer and a sequencing primer are designed via PyroMark Assay Design software according to a conserved region in a staphylococcus aureus plasma-coagulase DNA base sequence; and then a method for detecting staphylococcus aureus from the level of the DNA base sequence is established through a PCR-pyrophosphate sequencing method. In the invention, PCR augmentation is firstly carried out on a target segment under the condition of ensuring the specificity of the augmented segment; then a single-chain template is prepared from the augmented product; and then pyrophosphate sequencing is carried out under the guide of the sequencing primer. The base sequences obtained behind the plasma-coagulase DNA sequencing primer are discovered to have good specificity through repeated comparison by using a BLAST function of an NCBI (National Center of Biotechnology Information) website; and the staphylococcus aureus can be identified when 29 DNA base sequences behind the sequencing primer are CAACCGACGACACCGAACCCTATTTTAGA.

Description

technical field [0001] The invention belongs to the technical field of bioengineering, and in particular relates to detection primers, a kit and a detection method for detecting Staphylococcus aureus by PCR-pyrophosphate method. Background technique: [0002] Staphylococcus aureus (Staphylococcus aureus) is a major pathogen that causes nosocomial infection and bacterial food poisoning. % of the population is the carrier of the pathogen, Japan's survey results show that an average of 32.5% of the food there is Staphylococcus aureus contamination. With the development of molecular biology, there have been reports on the detection of Staphylococcus aureus by ordinary PCR or fluorescent PCR. Although these methods can achieve rapid and high-throughput purposes, they cannot obtain the gold standard of molecular diagnosis-DNA base sequence , so there is a possibility of false positives. Pyrosequencing (pyrosequencing) technology is a real-time short-segment DNA sequencing techno...

AI Technical Summary

Problems solved by technology

With the development of molecular biology, there have been reports on the detection of Staphylococcus aureus by ordinary PCR or fluorescent PCR. Although these methods can achieve rapid and high-throughput purposes, they cannot obtain the gold standard of molecular diagnosis-DNA base sequence , so there is a possibility of false positive

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