作　　者： ; ; ;

机构地区： 中国人民解放军第一军医大学热带军队卫生学系

出　　处： 《中国应用生理学杂志》 2004年第2期205-208,共4页

摘　　要： 目的 :探讨采用地高辛 (digoxin ,DIG)标记探针替代PCR Selectdifferentialscreening试剂盒中同位素标记探针的方法。方法 :抑制性消减杂交 (suppressionsubtractivehybridization ,SSH)所分离的差异表达序列转移至硝酸纤维素膜 ,以PCR法掺入DIG 11 dUTP制备探针 ,按DIG标记探针常规操作进行杂交显色 ,杂交阳性结果采用reverseNorthernblot再度杂交验证。结果 :应用DIG标记探针改良PCR SelectDifferentialScreening试剂盒所得到的阳性结果经reverseNorthernblot验证符合率达到 90 %。结论 :DIG标记探针改良PCR Selectdifferentialscreening试剂盒在避免了放射污染同时具备很高的特异性 ,完全可以替代同位素标记探针。 Aim: To explore a method to replace the isotope probe in the PCR-Select differential screening Kit with DIG labeled probe. Methods: Differential expressed sequences isolated from Suppression Subtractive Hybridization(SSH) was translocated onto nitric fibrin film. Probes were prepared with DIG-11-dUTP mixed in by PCR. Hybridization and coloration followed routine operation procedures of DIG lableled probe. Positive hybridization results were verified with reverse Northern blot. Results: The positive results from the PCR-Select differential screening Kit improved with DIG labeled probe achieved 90% correspondence verified by reverse Nor- thern blot. Conclusion: The PCR-Select differential screening Kit improved with DIG labeled probe maintaines high specificity while avoiding isotope pollution. It can replace isotope probe completely.

关 键 词： 抑制性消减杂交 地高辛 标记探针 同位素

领　　域： [生物学] [生物学]