PRDM14 and BLIMP1 control the development of chicken primordial germ cells

Primordial germ cell‐specific expression of eGFP in transgenic chickens

“…Thus, all of the PGC clones obtained were heterozygous gene-trapped cells. The lack of homozygous gene-trapped clones was consistent with our previous findings showing that PRDM14 was required for the proliferation of PGCs in vivo (Okuzaki et al, 2019). eGFP-positive PGCs purified by flow cytometry (not cloned) were transplanted into developing 2.5-day embryos as a preliminary trial.…”

“…We previously demonstrated that PRDM14 was essential for the proliferation of PGCs after the blastoderm because the number of PGCs was decreased by in vivo injection of replication-competent retrovirus vector containing shRNAs against PRDM14 (Okuzaki et al, 2019). We failed to observe that PRDM14 knockout decreased PGCs because of the early embryonic lethality of PRDM14 eGFP/eGFP .…”

“…This was unexpected because mice lacking Prdm14 were previously reported to be sterile, but viable (Yamaji et al, 2008), and the knockout of PRDM14 in Zebrafish did not cause early embryonic lethality, but resulted in motor neuron maturational arrest (Liu, Ma, Su, & Zhang, 2012). We previously showed that PRDM14 was expressed in cells other than PGCs, such as in the blastoderm (Okuzaki et al, 2019), and the loss of PRDM14 expression in these cells may have caused early embryonic lethality in PRDM14 eGFP/eGFP chickens. The mechanisms of the lethality, such as perturbed expression of pluripotency-related genes, are remained to be elucidated.…”

“…To clarify the function of PRDM14 using gene-trapped chickens, we designed a genome-editing strategy. CRISPR/Cas9-mediated homologous recombination was used to target the PRDM14 locus; the eGFP gene containing the stop codon was inserted immediately downstream of the initiation codon of PRDM14 based on the sequence we previously reported (Okuzaki et al, 2019). The targeting vector had a 797-bp sequence containing a 5 0 -untranslated region plus a coding sequence of 12 nucleotides as the 5 0 -homology arm, and a 779-bp sequence that contained a part of Exon 1 and the entire sequence from Intron 1 to Exon 3 as the 3 0 -homology arm.…”

“…We previously reported that the higher expression of PRDM14 was specific to PGCs, except for pluripotent cells at the blastodermal stage and neural plate cells at Hamburger-Hamilton stage (HH-stage) 4, as well as to the adult testis. Furthermore, PRDM14 was essential for the proper development of PGCs in vivo (Okuzaki et al, 2019). However, the role of chicken PRDM14 in early embryonic development remains unknown.…”