Human herpesvirus 6 (HHV-6) is a T-cell-tropic betaherpesvirus. HHV-6 can be classified into two variants, HHV-6 variant A (HHV-6A) and HHV-6B, based on genetic, antigenic, and cell tropisms, although the homology of their entire genomic sequences is nearly 90%. The HHV-6A glycoprotein complex gH/gL/gQ1/gQ2 is a viral ligand that binds to the cellular receptor human CD46. Because gH has 94.3% amino acid identity between the variants, here we examined whether gH from one variant could complement its loss in the other. Recently, we successfully reconstituted HHV-6A from its cloned genome in a bacterial artificial chromosome (BAC) (rHHV-6ABAC). Using this system, we constructed HHV-6ABAC DNA containing the HHV-6B gH (BgH) gene instead of the HHV-6A gH (AgH) gene in Escherichia coli. Recombinant HHV-6ABAC expressing BgH (rHHV-6ABAC-BgH) was successfully reconstituted. In addition, a monoclonal antibody that blocks HHV-6B but not HHV-6A infection neutralized rHHV-6ABAC-BgH but not rHHV-6ABAC. These results indicate that HHV-6B gH can complement the function of HHV-6A gH in the viral infectious cycle.