Fig. 6. In vitro transcription/translation of human nesprin-1 cDNA resulted in a 112 kDa product (lane 1). Translation reactions were as follows: nesprin-1 cDNA (lanes 1-9) and controls S. cerevisiae -factor (10-11) and ß-lactamase cDNA (12-13); either in the absence (1-2, 10,12) or in the presence of canine pancreatic microsome (3-9, 11,13); added either co-translationally (3-5, 11,13) or post-translationally (6-9). Reactions were digested with Proteinase-K alone (2, 4 and 7) or with 1% Triton X-100 (5). Translation products were subjected to sedimentation and separated into pellet (lane 8) and supernatant (lane 9) fractions. In these experiments, two controls were used to confirm microsome functionality. S. cerevisiae -factor exhibits an increase in molecular weight due to being N-glycosylated, and pro-ß-lactamase undergoes signal peptide cleavage to produce the lower molecular weight mature form.