原核移植技术对子代小鼠脑组织mRNA表达谱的影响

Study on expression profile of mRNA in brain of pronuclear transfer mice

目的探讨原核移植(PNT)技术对子代小鼠脑组织m RNA表达谱的影响。方法将SPF级ICR(CD-1)雌鼠超促排卵并与雄鼠交配受精后,收集雌鼠受精卵进行原核移植,获得重构胚胎,将其移植到假孕小鼠输卵管内获得原核移植子代小鼠(PNT组),未经原核移植操作的受精卵经过胚胎移植后获得子代小鼠(对照组)。提取2组小鼠脑组织RNA,逆转录为c DNA后荧光染料标记,利用Agilent小鼠m RNA芯片检测2组m RNA表达谱差异,筛选差异表达的m RNA并进行GO和信号通路分析。结果 PNT组与对照组表达差异倍数在2倍以上的m RNAs有392个,占所有m RNAs的1.7%,其中366个表达升高,26个表达降低;表达差异倍数在4倍以上的共11个。上述差异表达的m RNAs经GO分析结果显示靶基因富集于m RNA可变剪接、小GTP酶介导的信号转导、胰岛素受体信号通路调节等生物学过程以及水解酶活性、跨膜转运蛋白活性、焦磷酸酶活性等分子功能。信号通路富集分析结果显示靶基因集中在离子通道转运、脂肪酸代谢、丁酸甲酯代谢、三酰甘油和酮体代谢等信号通路。结论原核移植可能会对小鼠脑组织一些关键代谢过程产生影响。

Objective To investigate the expression profile of mRNAs in brain samples collected from pronuclear transfer(PNT) mice.Methods Female CD-1 mice were superovulated,and zygotes were collected after mating with adult male mice.Zygotes with two pronuclei were selected for pronuclear transfer manipulation,and then the reconstructed zygotes were transferred into the oviduct of pseudopregnant female mice.The infant mice obtained from pronuclear transfer were called PNT group,while the embryoes that were not performed pronuclear transfer was regarded as control group.Total RNA were extracted from brain samples of both PNT and control mice,and cDNA were labeled with fluorescent dye.Genes that were differentially expressed were identified using the Agilent mouse mRNA array.Gene ontology analysis and pathway analysis were also completed.Results Compared with control group,392 mRNAs were expressed differentially,which showed more than 2.0 times variation and statistical significance,accounting for 1.7%of all mRNAs.Among those 366 mRNAs were up-regulated and 26 mRNAs were down-regulated.Eleven mRNAs came to 4.0 times variation in total.Gene ontology analysis indicated that differentially expressed genes were significantly enriched in alternative mRNA splicing,small GTPase mediated signal transduction,regulation of insulin receptor signaling pathway,hydrolase activity,transmembrane transporter activity and pyrophosphatase activity.Significant enriched pathway terms contained ion channel transport,fatty acid metabolism,butanoate metabolism,triacylglycerol and ketone body metabolism.Conclusion Pronuclear transfer might influence some key metabolism process in mouse brain.