Construction and sequence analysis of subtraction complementary DNA libraries from human preimplantation embryos

Purpose: Because stage-specific genetic expression in human preimplantation development is not sufficiently studied, we have undertaken the constructi on of a subtraction complementary DNA (cDNA) library enriched for transcrip ts specific for human blastocysts. Methods: For this purpose individual pools of cDNAs synthesized from four h atched blastocysts and three cleaving 8- to 10-cell embryos were exposed to suppression subtractive hybridization to minimize the presence of transcri pts of housekeeping genes and other genes of maternal origin known to be ex pressed earlier in preimplantation development. Random clones of this libra ry were sequenced and analyzed using the BLAST algorithm. Results: The resulting subtraction library had a complexity of 3 x 10(5) an d an average size of inserts of about 0.8 kb. Sequencing of random library clones revealed the following human genes: CD9 antigen, fatty acid binding protein, ferritin heavy chain, amiloid precursor, MAP kinase messenger RNAs , DNA clone 127H14, messenger RNA for diacylglycerol kinase, a sequence hom ologous to C1 inhibitor; messenger RNA for the KIAA0145 gene, and others. Conclusions: The presence of these genes in human preimplantation developme nt suggests expression specific to the blastocyst stage.