Brady J P, Richmond R C, Oakeshott J G
Department of Biology, Indiana University.
Mol Biol Evol. 1990 Nov;7(6):525-46. doi: 10.1093/oxfordjournals.molbev.a040624.
A clone of the esterase-5 (Est-5) gene from Drosophila pseudoobscura has been isolated by hybridization to the cloned Est-6 gene of D. melanogaster. Southern analysis and sequencing of the cloned DNA revealed three regions of similarity to Est-6 that have been tentatively identified as genes, Est-5A, Est-5B, and Est-5C. Introduction of each of the three genes separately into D. melanogaster by P-element transformation has demonstrated that Est-5B encodes an enzyme with the same physical properties as EST 5 in D. pseudoobscura. Sequence analysis indicates that Est-5B encodes a 545-amino-acid protein and is composed of two exons separated by a 55-bp intron in the same position as the 51-bp intron in Est-6. Comparison of the Est-5B coding region with that of Est-6 reveals an overall similarity (73% at both the nucleotide and amino acid levels) that is substantially lower than that for other genes sequenced in both of these species. Total nucleotide and nonsynonymous site differences between Est-6 and Est-5B are more abundant in the second exon than in the first, suggesting differential effects of selection or mutation on these two exons. Comparisons of the 5'-flanking DNA of Est-5B and Est-6 reveal four short conserved sequence elements, but the remaining upstream sequences show no significant similarity. Conservation in the 3'-flanking DNA is limited to the presence of two polyadenylation sites that may correlate with the existence of two transcripts from both Est-5B and Est-6. The patterns of nucleotide substitutions and amino acid replacements between Est-5B and Est-6 are consistent with the hypothesis that mutation and genetic drift are responsible for the differences between these two genes.
通过与黑腹果蝇的酯酶-6（Est-6）基因克隆进行杂交，分离出了拟暗果蝇酯酶-5（Est-5）基因的一个克隆。对克隆的DNA进行Southern分析和测序，发现与Est-6有三个相似区域，暂定为基因Est-5A、Est-5B和Est-5C。通过P元素转化将这三个基因分别导入黑腹果蝇，结果表明Est-5B编码一种与拟暗果蝇中EST 5具有相同物理性质的酶。序列分析表明，Est-5B编码一个含545个氨基酸的蛋白质，由两个外显子组成，中间被一个55bp的内含子隔开，该内含子的位置与Est-6中的51bp内含子相同。将Est-5B的编码区与Est-6的编码区进行比较，发现总体相似度（核苷酸和氨基酸水平均为73%）明显低于这两个物种中测序的其他基因。Est-6和Est-5B之间的总核苷酸和非同义位点差异在第二个外显子中比在第一个外显子中更丰富，这表明选择或突变对这两个外显子有不同的影响。对Est-5B和Est-6的5'侧翼DNA进行比较，发现有四个短的保守序列元件，但其余上游序列没有明显的相似性。3'侧翼DNA的保守性仅限于存在两个聚腺苷酸化位点，这可能与Est-5B和Est-6的两个转录本的存在有关。Est-5B和Est-6之间的核苷酸替换和氨基酸替代模式与突变和遗传漂变导致这两个基因差异的假设一致。
