Murphy M H, Baralle F E
Nucleic Acids Res. 1983 Nov 25;11(22):7695-700. doi: 10.1093/nar/11.22.7695.
The transcription of tRNA and Alu repeat genes in vitro by RNA polymerase III has been shown to be dependent on the presence of two intragenic regions, which contain the consensus sequences RGYNNRRYGG (box A) and GA/TTCRANNC (box B), located 30-60 nucleotides apart. The role of box B and some of its variants was analyzed by a novel method involving the chemical synthesis of double stranded analogues of box B which were subsequently cloned into recombinant vectors carrying box A alone. This method creates a series of semi-synthetic RNA polymerase III promoters and has no limitation on the structure and number of variants which can be generated. The results showed the "wild type" sequence GTTCGAGAC and the sequence GTTCGTGAC (an A to T transversion of the 6th position) were active in promoting RNA polIII transcription. However, the box B sequences CTTCGAGAC and GTACGAGA, where the only departures from the consensus are a G to C and an A to T transversion in the 1st and 3rd positions respectively, were unable to restore promoter function.
体外实验表明，RNA聚合酶III对tRNA和Alu重复基因的转录依赖于两个基因内区域的存在，这两个区域包含位于30至60个核苷酸间距处的共有序列RGYNNRRYGG（A框）和GA/TTCRANNC（B框）。通过一种新方法分析了B框及其一些变体的作用，该方法涉及化学合成B框的双链类似物，随后将其克隆到仅携带A框的重组载体中。这种方法创建了一系列半合成RNA聚合酶III启动子，并且对可产生的变体的结构和数量没有限制。结果显示，“野生型”序列GTTCGAGAC和序列GTTCGTGAC（第6位的A到T颠换）在促进RNA polIII转录方面具有活性。然而，B框序列CTTCGAGAC和GTACGAGA（分别在第1位和第3位与共有序列的唯一差异是G到C和A到T颠换）无法恢复启动子功能。
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