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Transcriptional activity of ACL recombinants normalized by Renilla was significant difference with pGL3-Basic expect for construct -27 and -15 (P<0.01). Construct -919 contains highest activity. 3 times reduction of transcriptional activity from-919 to -679bp indicated that negative regulation factors located probably in this region. The activity started on construct -73 suggested the basal promoter activity was located within the -73 to +77bp region; Transcriptional activity of IDHβrecombinants was not significantly different between the recombinant -58 and pGL3-Basic. The activity started on construct -82, decreasing with the length of the fragment up to -164 in despite of a bit of fluctuation, and kept increasing from construct -164 up to -279. Thus, the basal promoter activity was located within the -82 to +16bp region, whereas the upstream 197bp conferred maximal transcriptional activity.

采用5'端缺失策略,结合启动子预测结果,分别构建了8个ACL基因和19个IDH3β基因启动子区重组子,将其转染猪PK15细胞系,并利用荧光素酶双报告基因系统检测了它们的活性,结果表明:在所构建的猪ACL基因5侧翼序列的8个重组子中,除pGL-ACL27和pGL-ACL15外,其它重组子的荧光素酶活性与阴性对照均达到极显著水平(P<0.01),表明它们均具有启动子活性,pGL-ACL919活性最强,到pGL-ACL679活性下降了将近3倍,说明从-919bp到-679bp区域可能存在负的调控位点,从pGL-ACL621到pGL-ACL73活性有小幅度下降,到pGL-ACL27活性降到与阴性对照水平无显著差异,表明维持该启动子的基本活性区域位于-73bp到+77bp之间;IDH3β基因启动子活性开始于pGL-IB82,然后虽然有些波动,但是活性一直下降直到-164bp处,之后活性又开始升高,直到-279bp处活性达到最高,这些表明维持该启动子的基本活性区域位于-82bp到+16bp之间,从-82到-279之间的179bp区域具有最高的启动子活性。

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