We observed efficient fiber trimerization, virus production and fiber incorporation into virus particles for the genomically modified viruses with 5T/41sSK fibers. A previous study showed for genomically fiber modified viruses expression, but no incorporation of Piperacillin Sodium HAdV-41 short fibers with peptide inserted into the AB, CD, or HI loops, or G region when a second HAdV-7 fiber was present. Of note, both the specific Lithocholic acid insertion positions and the inserted peptides differed from those in our study. Furthermore, the fiber tail domain was derived from HAdV-41, which we previously reported to result in reduced fiber trimerization and incorporation when compared with the native HAdV-5 tail used by us. Still, in the Schoggins study the short HAdV-41 fiber without peptide insertion was preferentially incorporated into viral particles in presence of the HAdV-7 fiber indicating that the peptide insertions affected fiber incorporation. We conclude that the EG, HI and IJ loops of the HAdV-41 short fiber knob are recommended for insertion of ligand peptides, establishing a panel of fiber scaffolds for this purpose. However, fiber trimerization and incorporation into virus particles need to be evaluated individually for each ligand. The Schoggins paper also reports maturation and cell entry defects for virus particles containing the HAdV-41 short fibers with insertions in the G region. Note that these viruses differ from our viruses also by deletion of the penton RGD motif, which might contribute to virus cell entry defects. Furthermore, maturation defects were dependent on the sequence of the inserted peptide. Although we did not explicitly investigate virus particle maturation, we were successful in producing high titer viruses that show efficient fiber incorporation and transduction similar or even superior to matching viruses containing the HAdV5 fiber. These results argue that the Ad5T/41sSK fiber scaffold described here and in our previous study is compatible with genomic fiber modification, facilitating the development of entry-targeted Ad vectors at high quantity and quality and of entry-targeted oncolytic Ads per se. Our study further demonstrates that entry targeting is not enhanced by replacement of the short HAdV-41-derived shaft in the chimeric Ad5T/41sSK fiber with the long HAdV-5 shaft, a strategy we pursued hypothesizing that ligand-receptor interactions would be improved.