Detection Of The Expression Levels Of MiR-21in Serum By Establishing SYBR Green Ⅰ FQ-PCR And Its Clinical Application In Different Cervical Diseases

Objective：To establish a method of SYBR Green I real-time quantitative PCR with stem-loop RTprimer for the detection of the expression levels of miR-21in serum. Then applicating themethod for the detection of the expression levels of miR-21in serum of different cervicaldiseases, to evaluate whether the levels of miR-21in serum can be used as a genemarker for theearly diagnosis and prognosis of cervical cancer.Methods：1. Total RNAs were extracted by the Trizol using the growth period of culture Hela cells. Thestem-loop reverse transcriptional primer, the primers of real-time PCR of miR-21had beendesigned and the expression levels of miR-21in cells or serum were detected with RT-PCRand FQ-PCR. The melting curve was analyzed to determine the specificity and stability ofthe expression.2. The method was used in the detection of the expression levels of miR-21in serum frompatients with different cervical diseases: serum samples were collected from15patients ofchronic cervicitis, uterine fibroids, cervical cancer and postoperative patients. With U6asthe internal control, we analyzed whether the expressions of miR-21in serum with differentcervical diseases were different, and the effects of miR-21expression before and afteroperation on prognosis of cervical cancer. All the data were analyzed by spss13.0, anda=0.05was regarded as the statistical criterion.Results：1. The method of SYBR Green I real-time quantitative PCR for the detection of the expressionlevels of miR-21in serum from patients with different cervical diseases was established. Themelting curve had shown a single peak, and the product of PCR was specific.2. The high expression levels of miR-21had been detected in serum of cervical cancerpatients，compared with that of the uterine fibroids and cervicitis, Meanwhile, the expressionlevels of miR-21has obviously decreased after surgical resection on cervical cancer patients(P<0.05). And that of cervical cancer patients after surgical resection decreasedobviously (P<0.05).The expression levels of miR-21in serum associated with cervicalcancer, and can be used for prognostic judgment, except for early diagnosis.Conclusions：The method of SYBR GreenⅠFQ-PCR with stem-loop RT primer is a rapid, sensitive, andspecific way to detect miRNA in serum, which can be a genemarker in early diagnosis andprognostic judgment of cervical cancer.