Owczarek C M, Enriquez-Harris P, Proudfoot N J
Sir William Dunn School of Pathology, University of Oxford, UK.
Nucleic Acids Res. 1992 Feb 25;20(4):851-8. doi: 10.1093/nar/20.4.851.
We have set up an experimental system to map the primary transcription unit of the human alpha 2 globin gene. The duplicated human alpha globin genes (alpha 2-alpha 1) were linked to the alpha globin locus Positive Regulatory Element (PRE) and stably transfected into murine erythroleukaemia cells. We then developed a quantitative reverse transcriptase, polymerase chain reaction assay to map alpha 2 primary transcripts using primer pairs derived from different parts of the alpha 2 globin gene and its 3' flanking region. This approach has revealed the presence of steady state nuclear RNA past the poly(A) site of the alpha 2 globin gene at approximately 40% of the level of unspliced intron transcript. Furthermore, these 3' flanking transcripts diminish 500 bp into the 3' flanking region, identifying this part of the alpha 2 globin gene as the principal region of termination of transcription.
我们建立了一个实验系统来定位人类α2珠蛋白基因的初级转录单元。将重复的人类α珠蛋白基因（α2-α1）与α珠蛋白基因座正调控元件（PRE）相连，并稳定转染到小鼠红白血病细胞中。然后，我们开发了一种定量逆转录聚合酶链反应测定法，使用源自α2珠蛋白基因及其3'侧翼区域不同部分的引物对来定位α2初级转录本。这种方法揭示了在α2珠蛋白基因的聚腺苷酸化位点之后存在稳态核RNA，其水平约为未剪接内含子转录本水平的40%。此外，这些3'侧翼转录本在进入3'侧翼区域500 bp后减少，确定α2珠蛋白基因的这一部分为转录终止的主要区域。
