The 27th immunoglobulin-like (I27) domain of muscle protein titin (schematically represented in Fig. 1D) was PCR amplified with sequence-specific primers containing PasI-modified ends and cloned so that one, two, and four copies of I27 (22 (link)) were obtained. The I27 domains were unidirectionally cloned into a unique PasI site (residue 29) of d37-41gp42 in the pCAGGS plasmid. The linkers were cloned just downstream of the transmembrane domain which ends at residue 22 and upstream of the gHgL binding domain which begins at residue 44. Other functional regions of gp42 were not disturbed. The ligated products were transformed into competent Escherichia coli DH5α and selected on plates containing ampicillin. DNA was isolated from overnight cultures using the Qiagen miniprep kit, digested to confirm the presence of the insert and correct orientation, and sequenced with primers internal and adjacent to the site of insertion and in both directions by the Northwestern Genomic Core Facility to confirm the resulting sequences. The resulting linker insertion sequences are shown in Table 1. Large-scale DNA preparations were isolated using Qiagen Endo-Free plasmid maxiprep kit and used in subsequent experiments. EBV gL in pCAGGS was previously described (28 (link)).