利用可誘導式調控系統影響斑馬魚生殖母細胞的遷移

Abstract

在斑馬魚胚胎發育4 cells至1000 cells時，生殖母細胞(Primordial Germ Cells, PGCs)在胚胎裡形成4群，之後會經由遷移，到達將來成體生殖腺發育的位置，經減數分裂產生配子，負責把親代的遺傳訊息傳遞到下一代。本實驗構築了pZBrBISRK質體。將此構築好的質體，以BstBI線性化，經顯微注射後，篩選出5株轉殖魚品系，並挑選了其中遺傳率(~45%)較高的ZBrBISRK2轉殖品系。在ZBrBISRK2的F2胚胎，在30 hpf時，在個體的頭部、軀幹以及尾部可以偵測到異位性nanos mRNA的表現，然而在野生型控制組中，nanos mRNA僅僅在gonad的位置有表現。進一步地，在Western blot的實驗中，能夠偵測到下游質體蛋白質專一的訊號。藉由以上的證據可知，利用誘導式調控系統確實造成PGCs在遷移時受到影響而產生異位性的遷移。

In embryogenesis of zebrafish, primordial germ cells (PGCs) formed four clumps before 1000-cell stage. PGCs would migrate and form two bilateral rows of cells close to the future gonad site. We micro-injected with a BstBI-cut pZBrBISRK plasmid and generated 5 transgenic line. One of which, ZBrBISRK2, having 45 % heredity rate, the highest among 5 transgenic lines, was selected. The nanos mRNA was detected not only in future gonad but also in head, trunk, and tail in the F2 embryos at 30 hpf derived from transgenic line ZBrBISRK2, whereas nanos mRNA was only detected in the gonad of wild type embryos, suggesting there were the mis-migrated PGCs. In Western blot, we could detect the positive bands in the total proteins extracted from ZBrBISRK2 F2 embryos. Therefore, we conclude that using conditionally regulatory system results in interfering the PGCs migration.