筛选代表小鼠植入前胚胎紧密化相关基因的EST Identification of ESTs of Genes Related to Compaction in Preimplantation Embryos of Mouse

分别收集181及241枚昆明白小鼠8细胞早期胚胎及8细胞紧密化胚胎，采用SMART PCR方法直接合成胚胎双链cDNA。进而运用抑制消减杂交技术（SSH）对8细胞早期胚胎及8细胞紧密化胚胎的基因表达进行研究，并将所获得的差异表达产物按片段大小分段分离纯化后克隆入pUCm-T载体中，经PCR鉴定后挑选阳性克隆进行测序，筛选出27个代表8细胞早期胚胎和紧密化8细胞胚胎差别表达基因的cDNA片段；经与GenBank中收录的序列进行同源性匹配分析，证实其中17个cDNA片段为新的EST，提交GeneBank后被接受并给予了新序列编号。这17个片段均可能为与紧密化密切相关的新基因的表达片段，为进一步克隆新的紧密化相关基因的全长cDNA及后续新基因的结构和功能研究打下基础。通过采用不同长度大小片段分别克隆的方法，可获得较长片段的EST，避免差异表达大片段的丢失。Abstract: A total of 181 8-cell embryos and 241 8-cell compacted embryos were collected respectively from Kunmingbai mouse and their cDNA was synthesized directly using SMART PCR. Genes, which expressed differently between early 8-cell embryos and 8-cell compacted embryos, were investigated using the method of suppression subtractive hybridization (SSH). Then PCR production was cloned into pUCm-T vector respectively according to the size after isolated and purified. Twenty-seven ESTs (expressed sequence tags ) of genes expressed differently between early 8-cell embryos and 8-cell compacted embryos have been isola