Guinta D R, Korn L J
Mol Cell Biol. 1986 Jul;6(7):2536-42. doi: 10.1128/mcb.6.7.2536-2542.1986.
In Xenopus laevis there are two multigene families of 5S RNA genes: the oocyte-type 5S RNA genes which are expressed only in oocytes and the somatic-type 5S RNA genes which are expressed throughout development. The Xenopus 5S RNA replication-expression model of Gottesfeld and Bloomer (Cell 28:781-791, 1982) and Wormington et al. (Cold Spring Harbor Symp. Quant. Biol. 47:879-884, 1983) predicts that the somatic-type 5S RNA genes replicate earlier in the cell cycle than do the oocyte-type genes. Hence, the somatic-type 5S RNA genes have a competitive advantage in binding the transcription factor TFIIIA in somatic cells and are thereby expressed to the exclusion of the oocyte-type genes. To test the replication-expression model, we determined the order of replication of the oocyte- and somatic-type 5S RNA genes. Xenopus cells were labeled with bromodeoxyuridine, stained for DNA content, and then sorted into fractions of S phase by using a fluorescence-activated cell sorter. The newly replicated DNA containing bromodeoxyuridine was separated from the lighter, unreplicated DNA by equilibrium centrifugation and was hybridized with DNA probes specific for the oocyte- and somatic-type 5S RNA genes. In this way we found that the somatic-type 5S RNA genes replicate early in S phase, whereas the oocyte-type 5S RNA genes replicate late in S phase, demonstrating a key aspect of the replication-expression model.
在非洲爪蟾中有两个5S RNA基因的多基因家族：仅在卵母细胞中表达的卵母细胞型5S RNA基因和在整个发育过程中都表达的体细胞型5S RNA基因。戈特斯费尔德和布卢默（《细胞》28:781 - 791，1982年）以及沃明顿等人（《冷泉港定量生物学研讨会》47:879 - 884，1983年）提出的非洲爪蟾5S RNA复制 - 表达模型预测，体细胞型5S RNA基因在细胞周期中比卵母细胞型基因更早复制。因此，体细胞型5S RNA基因在体细胞中结合转录因子TFIIIA方面具有竞争优势，从而得以表达，而卵母细胞型基因则被排除在外。为了验证复制 - 表达模型，我们确定了卵母细胞型和体细胞型5S RNA基因的复制顺序。用溴脱氧尿苷标记非洲爪蟾细胞，对DNA含量进行染色，然后使用荧光激活细胞分选仪将其分选到S期的各个部分。通过平衡离心将含有溴脱氧尿苷的新复制DNA与较轻的未复制DNA分离，并与针对卵母细胞型和体细胞型5S RNA基因的DNA探针杂交。通过这种方式，我们发现体细胞型5S RNA基因在S期早期复制，而卵母细胞型5S RNA基因在S期晚期复制，这证明了复制 - 表达模型的一个关键方面。
