论文部分内容阅读
目的:测定及分析GT198及其选择性剪接异构体GT198acDNA全序列并分别进行克隆,检测GT198a的功能并与GT198相比较。方法:RT-PCR及5′RACE测定GT198及GT198a的cDNA全序列;荧光素酶活性实验测定GT198a对基因转录的调控作用;免疫荧光实验检测GT198a对Rad51核聚体形成及GT198定位的影响。结果:GT198比GT198a上游多出100bp,与GT198相比,GT198a的序列包含了第一、二外显子间内含子。GT198a蛋白可使荧光素强度增加,Rad51核聚点形成受阻;而GT198蛋白则使荧光素强度减弱,对Rad51核聚点形成无明显影响。大量GT198a蛋白表达后GT198蛋白由核表达转为主要在胞质表达。结论:GT198a与GT198具有不同的5′端序列且在功能上相拮抗。GT198a可激活转录并抑制Rad51核聚点形成。GT198a蛋白的大量表达可促进GT198蛋白定位由胞核转向胞质。
OBJECTIVE: To determine and analyze GT198 and its alternative splicing isoforms GT198ac DNA sequence and cloned separately, to test the function of GT198a and GT198 compared. METHODS: The full-length cDNA of GT198 and GT198a was determined by RT-PCR and 5’RACE. The transcriptional regulation of GT198a by GT198a was determined by luciferase activity assay. The effect of GT198a on the formation of Rad51 nucleosomes and the localization of GT198 was detected by immunofluorescence assay. Results: GT198 was 100bp more upstream than GT198a. Compared with GT198, GT198a contained the first and second exon introns. GT198a protein can increase the intensity of fluorescein, Rad51 nuclear spot formation blocked; GT198 protein is weakening of the luciferin, Rad51 nuclear spot formation no significant effect. After a large number of GT198a protein expression GT198 protein from nuclear expression to the main expression in the cytoplasm. CONCLUSION: GT198a and GT198 have different 5 ’sequences and are functionally antagonistic. GT198a activates transcription and inhibits Rad51 nuclear spot formation. The high expression of GT198a protein can promote the localization of GT198 protein from nucleus to cytoplasm.