completely post translationally modified protein synthesis was carried out in the presence of the co and post translational modification machinery including glycosylation machinery besides the components of cell free protein synthesis the reaction mixture contained about 53 v v nuclease treated rrl and the final concentrations of other key components were 17 mm creatine phosphate 48 u03bcg ml creatine phosphokinase 40 u03bcm each amino acid 260 units ml sp6 rna polymerase 75 u03bcg ml circular plasmid dna 1 8 mm atp 1 3 mm gtp 1 mm each of utp and ctp 50 mm potassium acetate 3 6 mm magnesium acetate 0 4 mm spermidine 4 mm hepes koh ph 7 3 1600 units ml ribonuclease inhibitor 2 7 mm dtt 9 5 u03bcm hemin and 57 u03bcg ml calf liver total trna mixture the reaction mixture was incubated for 60 min at 30 u00b0 c fig1 shows an autoradiogram of completely post translationally modified epo by the method disclosed in the present invention and unmodified epo that has been separated by sds polyacrylamide gel electrophoresis lane 1 shows epo produced by cell free protein synthesis without co and post translational modification machinery lane 2 shows epo produced by cell free completely post translationally modified protein synthesis in lane.