胆囊收缩素cDNA探针制备及其在癫痫发病机制研究中的应用

①目的　制备胆囊收缩素（ Ｃ Ｃ Ｋ）ｃ Ｄ Ｎ Ａ 探针，并以此探针检测遗传性听源性癫痫易感大鼠癫痫发作后脑内 Ｃ Ｃ Ｋｍ Ｒ Ｎ Ａ 表达。②方法　将含０．５ｋｂｐ Ｒａｔ Ｃ Ｃ Ｋｍ Ｒ Ｎ Ａ 的质粒 Ｐ Ｕ Ｃ１３ 扩增、回收和纯化，用随机引物标记法，得到地高辛精（ Ｄｉｇ）标记 Ｃ Ｃ Ｋｃ Ｄ Ｎ Ａ 探针。用原位杂交技术，对癫痫发作后大鼠海马和大脑皮层 Ｃ Ｃ Ｋｍ Ｒ Ｎ Ａ 表达进行研究。③结果　 Ｃ Ｃ Ｋｃ Ｄ Ｎ Ａ 探针标记效率为５０ｍ ｇ／ Ｌ；遗传性听源性癫痫易感大鼠癫痫发作后脑内 Ｃ Ｃ Ｋｍ Ｒ Ｎ Ａ 表达显著增加。④结论　本实验制备的 Ｄｉｇ Ｃ Ｃ Ｋｃ Ｄ Ｎ Ａ 探针，具有较好的特异性和灵敏度；原位杂交实验证实， Ｃ Ｃ Ｋ 参与了癫痫发作过程。

PURIFICATION OF CHOLECYSTOKININ cDNA PROBE AND ITS APPLICATION IN THE MECHANISM STUDY OF EPILEPSY

Objective\ To prepare cholecystokinin(CCK) cDNA probe and to explore CCK mRNA expression with the probe in hereditary audiogenic epilepsy susceptible rat brain after epilepsy seizure.\ Methods\ Plasmid PUC13 containing 0.5 kbp Rat CCK mRNA was amplified, retrieved and purified. CCK cDNA probe was labelled with digoxigenin by random priming. CCK mRNA expressions in rat hippocampus and cerebral cortex after epilepsy seizure were studied by in situ hybridization.\ Results\ The labelling rate of CCK cDNA probe was 50mg/L. The CCK mRNA expression was significantly increased after epilepsy seizure in hereditary audiogenic epilepsy susceptible rat.\ Conclusion\ Dig CCK cDNA probe obtained in this study had good specificity and sensitivity. In situ hybridization study proved that CCK was involved in the process of epilepsy seizure.