Developmental enhancement of intracytoplasmic sperm injection (ICSI)-generated quail embryos by phospholipase Cζ cRNA

Shusei Mizushima, Soichi Takagi, Tamao Ono, Yusuke Atsumi, Akira Tsukada, Noboru Saito, Kiyoshi Shimada

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The present study was conducted to improve the rate and stage of blastoderm and embryo development of quail oocytes by intracytoplasmic sperm injection (ICSI) with quail phospholipase Cζ (PLCζ) cRNA. Blastoderm development of quail oocytes cultured for 24 or 72 h were staged under a stereomicroscope. The blastoderms were stained with 4,6-diamidino-2-phenylindole (DAPI) for cell division progress. A quail PLCζ cDNA clone was isolated from sperm by RT-PCR. The 1978 bp nucleotide sequence contained an ORF encoding 637 amino acids of protein. The deduced quail PLCζ protein consists of EF-hand domain, X and Y catalytic domain and C2 domain, but lacked a plextrin-homology (PH) domain at the N-terminus. RT-PCR analysis revealed that PLCζ mRNA is expressed only in the testis. ICSI into a quail oocyte without PLCζ cRNA induced blastodermal development in 16% (4/25) of the oocytes which reached stages III-VI 24h after culture. In contrast, ICSI together with PLCζ cRNA (in 3 n/ at 60 μg/m/) into a quail oocyte induced blastoderm development more than 2 fold (36.8%, 7/19) with embryos reaching stages VI-VII. Furthermore, 72 h after culture ICSI into a quail oocyte without PLCζ cRNA induced embryo development in 15.4% (2/13) of the oocytes and reached stages V-VII. In contrast, ICSI together with PLCζ cRNA (in 3n/ at 60 μg/m/) into a quail oocyte induced 3 fold more embryo development (50%, 9/18) with embryos between stages VI and over X. The microinjection of PLCζ cRNA significantly improved the conventional ICSI method by enhancing the proportion and the stages of embryo development. Accordingly, the present PLCζ cRNA microinjection method in birds may contribute to assist in the production of transgenic birds and protection of endangered species of birds.

Original languageEnglish
Pages (from-to)152-158
Number of pages7
JournalJournal of Poultry Science
Issue number2
Publication statusPublished - May 15 2008
Externally publishedYes



  • Blastoderm
  • Embryo development
  • ICSI
  • Phospholipase Cζ
  • Quail

ASJC Scopus subject areas

  • Animal Science and Zoology

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