Doran J L, Bingle W H, Roy K L
Department of Microbiology, University of Alberta, Edmonton, Canada.
Gene. 1988 May 30;65(2):329-36. doi: 10.1016/0378-1119(88)90471-4.
Two human DNA fragments of 16.7 and 15.5 kb have been selected from a human lambda Charon-4A library by hybridization to an unfractionated tRNA probe. Restriction mapping and Southern and Northern hybridization analyses revealed the presence of a single tRNA-hybridizing region in each of the human DNA fragments. Nucleotide sequence analysis has identified two identical members of the tRNA(GCCGly) gene family. These tRNA(GCCGly) genes encode all of the conserved and semiconserved nucleotides of the tDNA split promoter sequences. Neither gene contains introns or encodes the CCA sequence present on the 3' terminus of mature tRNA. One of these identical tRNA(GCCGly) genes was found to be expressed at a substantially greater efficiency than the other in a HeLa cell lysate in vitro transcription system. No similarity was detected in the nucleotide sequences flanking these genes other than the characteristic, 3' oligo[dT] transcription termination signals and a TCTTT sequence located 7 to 10 bp upstream. These data are consistent with the hypothesis that as yet unidentified tDNA flanking sequences may have an important role in modulating human tRNA gene expression.
通过与未分级的tRNA探针杂交，从人λ噬菌体Charon - 4A文库中筛选出了两个分别为16.7 kb和15.5 kb的人类DNA片段。限制性图谱分析以及Southern和Northern杂交分析表明，这两个人类DNA片段中均存在单个与tRNA杂交的区域。核苷酸序列分析鉴定出了tRNA（GCCGly）基因家族的两个相同成员。这些tRNA（GCCGly）基因编码了tDNA分裂启动子序列中所有保守和半保守的核苷酸。这两个基因均不包含内含子，也不编码成熟tRNA 3'末端的CCA序列。在体外转录系统的HeLa细胞裂解物中，发现其中一个相同的tRNA（GCCGly）基因的表达效率明显高于另一个。除了特征性的3'寡聚[dT]转录终止信号和位于上游7至10 bp处的TCTTT序列外，在这些基因两侧的核苷酸序列中未检测到相似性。这些数据与以下假设一致：尚未鉴定的tDNA侧翼序列可能在调节人类tRNA基因表达中起重要作用。
