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目的应用实时荧光PCR技术,检测14-3-3β基因在新疆Kaposi肉瘤(Kaposi’s sarcoma,KS)中的表达并分析其与KS发生发展的关系。方法2005至2006年收集新疆部分地区的7例经典型KS患者的肿瘤和正常皮肤组织,并经病理活检证实为KS。以SYBRGreenⅠ为荧光指示剂,GAPDH作内参照,建立检测14-3-3βmRNA表达的实时荧光PCR反应体系。分析14-3-3βmRNA的表达与KS发生发展的关系。结果①通过对实验参数的优化,适宜的反应条件为退火温度60℃,循环次数45次;②KS组14-3-3βmRNA的表达明显高于正常对照组(P<0.05)。结论采用SybrGreenⅠ的Smartcycler系统的实时荧光PCR方法检测14-3-3βmRNA快速、简便、稳定性好。14-3-3βmRNA的表达与KS的发生发展有重要关系。
Objective To detect the expression of 14-3-3β gene in Kaposi’s sarcoma (KS) by real-time fluorescence PCR and analyze its relationship with the development of KS. Methods Tumors and normal skin tissues from 7 patients with classic KS in some areas of Xinjiang were collected from 2005 to 2006 and confirmed as KS by pathological biopsy. Using SYBR Green I as fluorescent indicator and GAPDH as internal reference, a real-time fluorescence PCR reaction system for detecting 14-3-3β mRNA expression was established. To analyze the relationship between the expression of 14-3-3β mRNA and the development of KS. Results (1) Through the optimization of the experimental parameters, the suitable reaction conditions were annealing temperature 60 ℃ and number of cycles 45 times. ② The expression of 14-3-3βmRNA in KS group was significantly higher than that in normal control group (P <0.05). Conclusion The real-time fluorescent PCR method using SybrGreen Ⅰ’s Smartcycler system can detect 14-3-3β mRNA rapidly, easily and stably. The expression of 14-3-3|ÂmRNA plays an important role in the development of KS.