Cloning Of Swine H1foo Gene And Preliminary Research Of Its Effection On Oocyte Meiotic Maturation

H1foo (oocyte-specific linker histone H1) is a linker histone specifically expressed inoocytes and early embryos in mammalian; it plays a crucial role in oogenesis, fertilizationand embryogenesis. The objectives of this study are to clone swine H1foo gene and examineits role in the process of the first meiosis.In the research, firstly, we applied the sequence information in NCBI and5′RACE toclone the H1foo gene of swine, and then the H1foo CDS was linked into pVenus to constructa eukaryotic expression vector pVenus-H1foo; Secondly, the expression variation of H1foomRNA in different stage of oocytes and embryos was examined through QRT-PCR; At last,the role of H1foo in the process of the first meiosis was examined through the way ofover-expression and interference.The results of the research were as follows:1. We obtained the complete CDS of swine H1foo gene by5′RACE and RT-PCR. Theresult of sequence analysis showed that the CDS region of the swine H1foo gene includes1041nucleotides, encoding for346amino acids(Mol.wt.:36.45kD); The nucleotide similarityof H1foo between swine and bovine, human, mice is75.7%,67.9%and54.3%, respectively;The similarity of H1foo protein between swine and bovine, human, mice is75.7%,64.8%and49.4%respectively, however, the similarity of the globular domain is95.7%,92.8%and82.6%respectively. The sequence information was submitted to GenBanK (GenBankaccession No.: HQ915640).2. The expression variation of H1foo mRNA in different stages of oocytes and embryoswas detected by QRT-PCR. In the result, the highest level of H1foo transcripts was in the MIIoocytes; The H1foo mRNA level of MII oocytes was more than one time higher than GVoocytes; There was a continuous decline through development of embryo afterparthenogenesis activation, the mRNA level compared to the amount of the MII oocytedeclined to23%,14.97%,11.24%and3.79%in the2-cells,4-cells,8-cells and16-cellsembryos, respectively, and it was almost undetectably in blastocyst.3. There was no effect on the PBEI (First Polar Body Extrusion) when we injected H1foo RNA in GV oocytes to over-expression. However, when the H1foo was interfered withantisense morpholino oligonucleotides, the percentage of PBEI declined by25%compared tothe control oocytes, and the major of the immaturate oocytes were stay in the pre-MI stage;Neither over-expression nor interference had effect on the spindle structure.Conclusion: In this study, we obtained the complete CDS of swine H1foo gene; TheH1foo mRNA level of MII oocytes was more than one time higher than GV oocytes, therewas a continuous decline through development of embryos after parthenogenesis activation;Our results indicated that down-regulation of H1foo impaired the process of the first meiosis.