Hirabayashi Masumi, Hara Hiromasa, Goto Teppei, Takizawa Akiko, Dwinell Melinda R, Yamanaka Takahiro, Hochi Shinichi, Nakauchi Hiromitsu
Center for Genetic Analysis of Behavior, National Institute for Physiological Sciences, Aichi 444-8787, Japan.
School of Life Science, The Graduate University for Advanced Studies, Aichi 444-8787, Japan.
J Reprod Dev. 2017 Dec 15;63(6):611-616. doi: 10.1262/jrd.2017-074. Epub 2017 Aug 19.
The present study was conducted to establish haploid embryonic stem (ES) cell lines using fluorescent marker-carrying rats. In the first series, 7 ES cell lines were established from 26 androgenetic haploid blastocysts. However, only 1 ES cell line (ahES-2) was found to contain haploid cells (1n = 20 + X) by fluorescence-activated cell sorting (FACS) and karyotypic analyses. No chimeras were detected among the 10 fetuses and 41 offspring derived from blastocyst injection with the FACS-purified haploid cells. In the second series, 2 ES cell lines containing haploid cells (13% in phES-1 and 1% in phES-2) were established from 2 parthenogenetic haploid blastocysts. Only the phES-2 cell population was purified by repeated FACS to obtain 33% haploid cells. Following blastocyst injection with the FACS-purified haploid cells, no chimera was observed among the 11 fetuses; however, 1 chimeric male was found among the 47 offspring. Although haploid rat ES cell lines can be established from both blastocyst sources, FACS purification may be necessary for maintenance and chimera production.
本研究旨在利用携带荧光标记的大鼠建立单倍体胚胎干细胞（ES）系。在第一个系列中，从26个孤雄单倍体囊胚中建立了7个ES细胞系。然而，通过荧光激活细胞分选（FACS）和核型分析，仅发现1个ES细胞系（ahES-2）含有单倍体细胞（1n = 20 + X）。在将FACS纯化的单倍体细胞注射到囊胚后获得的10个胎儿和41个后代中，未检测到嵌合体。在第二个系列中，从2个孤雌单倍体囊胚中建立了2个含有单倍体细胞的ES细胞系（phES-1中为13%，phES-2中为1%）。仅对phES-2细胞群体进行了反复的FACS纯化，以获得33%的单倍体细胞。将FACS纯化的单倍体细胞注射到囊胚后，在11个胎儿中未观察到嵌合体；然而，在47个后代中发现了1只嵌合雄性大鼠。尽管单倍体大鼠ES细胞系可以从两种囊胚来源建立，但FACS纯化对于维持细胞系和产生嵌合体可能是必要的。
