Expression of RNA from developmentally important genes in preimplantation bovine embryos produced in TCM supplemented with BSA.
Expression of RNA from developmentally important genes in preimplantation bovine embryos produced in TCM supplemented with BSA.
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在添加 BSA 的中药中产生的植入前牛胚胎中发育重要基因的 RNA 表达。
DOI：
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发表时间：
1998
期刊：
影响因子：
--
通讯作者：
Heiner Niemann
中科院分区：
文献类型：
--
作者：
C. Wrenzycki;D. Herrmann;J. Carnwath;Heiner Niemann
本研究研究了半限定培养系统对参与压缩和空化的基因 RNA 表达时间模式的影响：间隙连接蛋白 connexin43 (Cx43)、桥粒糖蛋白桥粒糖蛋白 1 (Dg 1)、桥粒胶蛋白 I、II 和 III (Dc I、Dc II、Dc III)、桥粒蛋白 plakophilin (Plako)；代谢葡萄糖转运蛋白-1 (Glut-1)； RNA 加工聚 (A) 聚合酶 (PolyA)；热休克蛋白70.1（HSP）；使用添加 BSA 的 TCM199 作为培养基体外生成的牛卵母细胞和胚胎中的滋养层功能滋养层蛋白 (TP)。从超排卵小母牛中收集体内衍生的桑葚和囊胚，也用于本研究。从 20-50 个卵母细胞或胚胎池中提取 Poly(A)+ RNA，通过逆转录聚合酶链式反应进行分析，并通过测序验证扩增的片段。每个发育阶段的测定至少重复三次，并在所有重复中提供一致的结果。在体外产生的牛胚胎中，直到桑椹胚阶段都可检测到编码Cx43的mRNA，而囊胚和孵化的囊胚不表达该基因。在整个测试的植入前阶段，没有发现 Dg 1 和 Dc I 的转录本。从 2-4 细胞胚胎直至孵化的囊胚阶段都发现了 Dc II 和 Dc III 转录本。在未成熟和成熟的卵母细胞和受精卵中检测到编码 Plako 的 mRNA，而在 2-4 细胞和 8-16 细胞胚胎中未观察到转录本。该基因从桑葚胚到孵化的囊胚阶段再次表达。体外产生的卵母细胞和牛胚胎在整个植入前发育直至孵化囊胚阶段都显示出 Glut-1、PolyA 和 HSP 的转录物。编码TP的基因仅在囊胚和孵化的囊胚中转录。体内产生的桑葚和囊胚与体外对应物表现出相同的表达，但有一个例外：体内胚胎转录 Cx43。这项研究的结果首次揭示了参与早期牛胚胎发育各个过程的一组“标记”基因的转录模式。体外产生的可转移桑葚和胚泡表达的大多数基因与体内对应物相似。这些数据有助于对这种广泛使用的牛胚胎体外培养系统进行分子表征，并为生产“生理正常”胚胎提供了重大进展。
This study investigated the effects of a semi-defined culture system on the temporal pattern of expression of RNA from genes involved in compaction and cavitation: gap junction protein connexin43 (Cx43), desmosomal glycoproteins desmoglein 1 (Dg 1), desmocollins I, II and III (Dc I, Dc II, Dc III), desmosomal protein plakophilin (Plako); metabolism glucosetransporter-1 (Glut-1); RNA processing poly(A)polymerase (PolyA); heat shock protein 70.1 (HSP); and trophoblastic function trophoblast protein (TP) in bovine oocytes and embryos generated in vitro using TCM199 supplemented with BSA as the culture medium. Morulae and blastocysts derived in vivo were collected from superovulated heifers and also used for this study. Poly(A)+ RNA was extracted from pools of 20-50 oocytes or embryos, analysed by reverse transcription-polymerase chain reaction and the amplified fragments were verified by sequencing. Assays were repeated at least three times for each developmental stage and provided consistent results in all replicates. In bovine embryos produced in vitro, mRNA encoding Cx43 was detectable up to the morula stage, whereas blastocysts and hatched blastocysts did not express this gene. No transcripts were found for Dg 1 and Dc I throughout the tested preimplantation stages. Dc II and Dc III transcripts were found from 2-4-cell embryos up to the hatched blastocyst stage. mRNA encoding Plako was detected in immature and mature oocytes and zygotes, while no transcripts were seen in 2-4-cell and 8-16-cell embryos. The gene was expressed again from the morulae to the hatched blastocyst stage. Oocytes and bovine embryos produced in vitro showed transcripts for Glut-1, PolyA and HSP throughout preimplantation development up to the hatched blastocyst stage. The gene encoding TP was transcribed only in blastocysts and hatched blastocysts. Morulae and blastocysts produced in vivo showed the same expression as their in vitro counterparts, with one exception: the in vivo embryos transcribed Cx43. The results of this study reveal for the first time the transcriptional pattern of a set of 'marker' genes involved in various processes in early bovine embryonic development. Transferable morulae and blastocysts produced in vitro expressed most genes similar to their in vivo counterparts. These data contribute to the molecular characterization of this widely used in vitro culture system for bovine embryos and provide a major advance towards production of 'physiologically normal' embryos.
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DOI：
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发表时间：
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期刊：
The Journal of biological chemistry
影响因子：
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Development (Cambridge, England)
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