Bodary S, Grossi G, Hagenbüchle O, Wellauer P K
J Mol Biol. 1985 Mar 5;182(1):1-10. doi: 10.1016/0022-2836(85)90022-1.
Several members of the Amy-2 alpha-amylase multigene family from the CE/J strain of mouse have been cloned in cosmid vectors. Structural analysis of these recombinants reveals that the cloned Amy-2 copies contain tandem 5' termini. The duplicated 5'-terminal elements, which lie upstream from the Amy-2 cap site, are separated from their Amy-2 homologues by about 8000 bases. The orientation of these 5' orphons is the same as that of Amy-2. Gene titration and cloning experiments suggest that at least four of the approximately 15 Amy-2 copies present in the CE/J genome contain 5' orphon elements. The extent of sequence homology between 5' orphons and their gene homologues has been determined by DNA sequence analysis. All the orphons are identical and contain the entire 185 base-pairs of the first exon, 49 base-pairs of the first intron and more than 400 base-pairs of the Amy-2 5' flanking region. Intron and flanking-region sequences of the orphons differ by about 20% from their Amy-2 counterparts, and the exon by about 8%. The TATA box and the cap site are conserved, while the ATG translation initiation signal is mutated to ATA in the orphon. No transcription initiation has been detected at the orphon cap site using run-off transcription in isolated pancreatic nuclei in vitro.
已从小鼠CE/J品系的Amy-2α-淀粉酶多基因家族中克隆出几个成员，并将其克隆到黏粒载体中。对这些重组体的结构分析表明，克隆的Amy-2拷贝含有串联的5'末端。位于Amy-2帽位点上游的重复5'末端元件与其Amy-2同源物相隔约8000个碱基。这些5'旁系同源序列的方向与Amy-2相同。基因滴定和克隆实验表明，CE/J基因组中约15个Amy-2拷贝中至少有4个含有5'旁系同源元件。通过DNA序列分析确定了5'旁系同源序列与其基因同源物之间的序列同源程度。所有旁系同源序列均相同，包含第一个外显子的完整185个碱基对、第一个内含子的49个碱基对以及Amy-2 5'侧翼区域的400多个碱基对。旁系同源序列的内含子和侧翼区域序列与其Amy-2对应序列相差约20%，外显子相差约8%。TATA盒和帽位点是保守的，而旁系同源序列中的ATG翻译起始信号突变为ATA。在体外分离的胰腺细胞核中使用径流转录法未在旁系同源序列的帽位点检测到转录起始。
