Yenikolopov G N, Kuzin B A, Evgen'ev N B, Ludwig M Z, Korochkin L I, Georgiev G P
Institute of Molecular Biology, USSR Academy of Sciences, Moscow.
EMBO J. 1983;2(1):1-7. doi: 10.1002/j.1460-2075.1983.tb01371.x.
We have cloned the gene for the esterase S isozymes complex from the genome of Drosophila virilis in pBR322. Esterase S is an enzyme which is specifically synthesized in the ejaculatory bulbs of D. virilis adult males. The gene for the esterase S isozyme complex (estS) has been localized in band 2G5e of chromosome II. Poly(A)+ RNA prepared from ejaculatory bulbs actively hybridizes with this band. A cloned 15-kb fragment of D. virilis DNA (pVE9) also hybridizes with band 2G5e. The area encoding the poly(A)+ RNA is located in the middle part of the cloned fragment whose ends are not transcribed in vivo. Only one poly(A)+ RNA which is 1.9 kb long and complementary to pVE9 DNA can be revealed in the cytoplasm. The mRNA preselected by hybridization to pVE9 DNA was microinjected into the cytoplasm of Xenopus laevis oocytes. In other experiments, the pVE9 DNA itself was microinjected into oocyte nuclei. In both cases, esterase S is synthesized in the oocytes, and the major part of the protein is transported from the oocytes and accumulated in the incubation medium.
我们已将果蝇(Drosophila virilis)基因组中酯酶S同工酶复合物的基因克隆到pBR322中。酯酶S是一种在成年雄性果蝇的射精球中特异性合成的酶。酯酶S同工酶复合物(estS)的基因已定位在二号染色体的2G5e带。从射精球制备的多聚腺苷酸加尾RNA(poly(A)+ RNA)与该带发生活跃杂交。一个克隆的15千碱基对(kb)的果蝇DNA片段(pVE9)也与2G5e带杂交。编码多聚腺苷酸加尾RNA的区域位于克隆片段的中部,其两端在体内不被转录。在细胞质中只能检测到一种长度为1.9 kb且与pVE9 DNA互补的多聚腺苷酸加尾RNA。通过与pVE9 DNA杂交预选的信使核糖核酸(mRNA)被显微注射到非洲爪蟾(Xenopus laevis)卵母细胞的细胞质中。在其他实验中,pVE9 DNA本身被显微注射到卵母细胞核中。在这两种情况下,卵母细胞中都会合成酯酶S,并且大部分蛋白质会从卵母细胞中转运出来并积累在培养液中。