Combined use of RT-PCR and gel electrophoresis to detect expression of transforming growth factor beta1 in mouse lung fibroblasts in vitro / 中国组织工程研究

BACKGROUND: As a combination of reverse transcription (RT) and polymerase chain reaction (PCR), RT-PCR has been used to detect gene expression levels in cells and tissues, RNA virus contents in cells and specific gene cloned cDNA sequences.OBJECTIVE: To detect the inhibitory effcet of Stealth siRNAs on the expression of transforming growth factor β1 (TGF-β1).METHODS: There were blank control, empty vector transfection, stealth-48, stealth-166, and stealth-594 groups. Three stealth siRNAs aimed at different sequences in TGF-β1 mRNA were made, and were then transfected into BALB/c mouse lung fibroblasts in vitro. The expressions of TGF-β1 and connective tissue growth factor were detected by RT-PCR.RESULTS AND CONCLUSION: In different time periods, the TGF-β1 expression was differentially depressed by three stealth siRNAs, especially stealth-166. The inhibitory effects varied with time, which could be detective at 48 hours,reached the peak at 72 hours and then began to attenuate at 96 hours. Our findings show that the inhibitory effect of stealth siRNAs on the TGF-β1 expression in mouse lung fibroblasts can be detected by RT-PCR.

Correlation analysis of positive rate of HPV genotyping test and HPV nucleic acid loads / 中华微生物学和免疫学杂志

Objective To investigate the correlation between the positive rate of PCR-reverse dot blot genotyping test and the loads of the viral nucleic acid. Methods The fluorescent PCR assay was used to detect the high-risk HPV(HR-HPV)DNA loads in the cervical mucus samples from 1162 female pa-tients. Patients with positive HR-HPV DNA were further analyzed by PCR-reverse dot blot hybridization as-say for HPV genotyping. Results The overall positive rate of genotyping test was 68. 8% . The positive rate of genotyping test had a significant positive correlation with the Log Koc of HR-HPV DNA loads(r=0. 944, P﹤0. 01). The quadratic curve fitting formula was Y= -1. 806+0. 558X-0. 031X2(Y for genotyping positive rate,X for Log Koc of HR-HPV DNA loads). There were significant differences with the positive rate of gen-otyping test among patients with different viral loads(P﹤0. 01). When HR-HPV DNA loads were 104-105 copies/ ml,105-106 copies/ ml,106-107 copies/ ml and more than 107 copies/ ml,the positive rate of HPV genotyping test were 27. 8% ,48. 5% ,74. 0% ,97. 5% and 33. 3% ,51. 5% ,78. 0% ,97. 5% respective-ly by using different genotyping detection reagents. Conclusion The positive rate of PCR-reverse dot blot genotyping test was correlated with the loads of HPV nucleic acid.