Differentially Expressed Genes In Pig 2-Cell Stage Parthenogenetic Embryos With Different Developmental Competence

First cleavage time of embryos is an important criterion to evaluate embryo quality and potential developmental competence. We select embryos with different viability as research objects based on their different first cleveage time. Then through some experimental techniques, we will find some maternal genes which affect embryos developmental competence. Embryonic genome activation (ZGA) of pig occurs at 4-cell stage. If the development before that are controlled by maternal mRNAs, the levels of these maternal mRNAs are most likely different between early cleavage embryos and late cleavage embryos at 2-cell stag. Identification of mRNAs can help us find out maternal mRNAs which are critical to early development of embryos, and may contribute to finding out why first cleveage time can affect embryo developmental competence.Our study obtained developmental competent and incompetent two-cell stage pig parthenogenetic embryos based on different first cleavage time and the level of their mRNAs was compared. Differential Display Reverse Transcription Polymerase Chain Reaction was carried out on mRNAs isolated from these embryos to find out genes that expressed differently. Then use Reverse Northern-blot to verify these genes. After that, pig CPEB2(E3) cDNA was cloned by PCR and 5'Rapid-Amplification of cDNA Ends and its expression pattern was obtained by SYBR-Green real-time PCR.1. Eight fragments (E1-E4; L1-L4) differentially expressed between the two groups by Reverse Northern-blot. Four fragments (E1-E4) have a higher expressed level in early cleavage embryos (good quality); four fragments (L1-L4) have a higher expressed level in late cleavage embryos (bad quality).2. After cloning, sequencing and comparing with the database of swine genome and transcripts on NCBI, six in eight positive fragments had homologue ESTs3. After comparing with the database of human genome and transcripts on NCBI, E3 has high homology (the homology is 97%) with human CPEB2; and L3 has high homology (the homology is 89%) with hnRNPA1.4. Cloned CPEB2 cDNA of pig for first time. Submitted it to NCBI and received the Genbank accession: HM037344.5. Ascertained that CPEB2 mRNA exist in pig oocytes and embryos. Through SYBR GREEN real-time PCR, we make sure once again that the abundance of E3 in early cleavage embryos are higher than late cleavage embryos, and its expression pattern is similar to maternal mRNAs.