Construction of EGFR gene G719S and T790M mutation vector and preliminary clinical application / 临床检验杂志

ABSTRACT

Objective To construct mutant recombinant vector of epidermal growth factor receptor ( EGFR) gene G719S and T790M sites associated with cervical cancer, lay the foundation for the detection of EGFR gene mutation in cervical cancer. And using it to es-tablish a molecular switch platform to detect cervical cancer EGFR gene mutations. Methods Using the wild-type recombinant plasmid as template, the mutant fusion target fragment were amplified by overlap PCR, then connect this target fragment into the vector pMD19-T. The constructed mutant recombinant plasmid was finally transformed into competent cells E.coli DH5αfurther identified by PCR with bacterial solution and genome sequencing. Establishing the molecular switch for the detection of clinical cervical cancer samples. Re-sults The G719S and T790M mutations were successfully certified by genome sequencing, and the site-directed mutant vector was successfully constructed. In addition, a molecular switch detection platform was also successfully established for the detection of cervical cancer tissue DNA. Conclusion We successfully constructed an EGFR gene mutant recombinant vector by overlap PCR technique, which providing a new technical means for gene site-directed mutagenesis. And the molecular switch detection platform was successfully established based on it, which furnishing a new method for clinical detection of EGFR gene mutations.