Method for high-pass multi-enrichment quantitative PCR (ME-qPCR) detection of transgenic crop element and strain

Abstract

The invention relates to molecular biology, and particularly discloses a method for detection of transgenic crops.Inner primers and outer primers are designed according to the endogenous gene, exogenous gene and transgenic strain flanking sequence of a transgenic crop, all the inner primers and outer primers are mixed firstly for the first round of high-pass multi-enrichment quantitative PCR with a small cycle number (15 cycles), then the second round of fluorogenic quantitative PCR is conducted with the inner primers, a result is analyzed according to an amplification curve and a fusion curve, and in this way, the success rate and stability of the method are improved remarkably and the purpose of high-pass detection of transgenic crops can be realized.The sensitivity of the method is higher than that of ordinary fluorogenic quantitative PCR by about one magnitude order; the method has the same quantitative capacity as ordinary fluorogenic quantitative PCR, has high specificity and is a more effective method for detection of transgenic crops.

Description

technical field [0001] The invention relates to molecular biology, in particular to a method for high-throughput detection of transgenic crops. Background technique [0002] In transgenic detection, nucleic acid-based detection methods are widely used due to their strong stability and high specificity, among which PCR-based detection methods are particularly important. PCR-based detection methods mainly include ordinary PCR and fluorescent quantitative PCR. Fluorescent quantitative PCR is widely used in actual detection because it does not require gel electrophoresis and has strong specificity. However, the detection throughput has always been a bottleneck of PCR-based detection methods. For example, multiplex PCR is easy to form primer dimers between primers, and there is easy competition between primers, which leads to low detection throughput of multiplex PCR. ; and multiplex fluorescent quantitative PCR is also difficult to achieve high throughput due to the limitations...

AI Technical Summary

Problems solved by technology

However, the detection throughput has always been a bottleneck of PCR-based detection methods. For example, multiplex PCR is easy to form primer dimers between primers, and there is easy competition between primers, which leads to low detection throughput of multiplex PCR. ; and multiplex fluorescent quantitative PCR is also difficult to achieve high throughput due to the limitation of instrument detection fluorescence pathway and the competition between primers and probes

Images