HLA-G limited polymorphic gene maps to the human major histocompatibility complex (MHC) class I subregion and encodes the molecule which is the only MHC class I antigen expressed on cytotrophoblast cells at the maternal-fetal interface. In this tissue, HLA-G primary mRNA is differentially spliced. We have used a sensitive hot start reverse transcriptase-polymerase chain reaction (RT-PCR) technique to investigate the expression of HLA-G gene in first trimester trophoblasts and adult peripheral blood cells. PCR amplification with HLA-G primers specific of exon 3 has enabled us to demonstrate a novel alternatively spliced form of HLA-G mRNA present in fetal first trimester trophoblasts and lacking exon 4 (HLA-G4). Cloning the whole PCR product and hybridizing recombinant bacterial colonies with specific probes has permitted evaluation of HLA-G4 vs. full length mRNA frequency at approximately 1:200. Moreover, the presence of HLA-G transcripts was found at a very weak level in adult peripheral blood lymphocytes and equally in B- and T-cell populations. These results are relevant in the context of immune tolerance and in the potential use of HLA-G transcripts as a marker for RT-PCR detection of the fetal cells in maternal as a marker for RT-PCR detection of the fetal cells in maternal blood.