The extreme strength and elasticity of spider silks originate from the modular nature of their repetitive proteins. To exploit suchmaterials and mimic spider silks, comprehensive strategies to produce and spin recombinant fibrous proteins are necessary. Thisprotocol describes silk gene design and cloning, protein expression in bacteria, recombinant protein purification and fiber formation.With an improved gene construction and cloning scheme, this technique is adaptable for the production of any repetitive fibrousproteins, and ensures the exact reproduction of native repeat sequences, analogs or chimeric versions. The proteins are solubilized in1,1,1,3,3,3-hexafluoro-2-propanol (HFIP) at 25-30% (wt/vol) for extrusion into fibers. This protocol, routinely used to spin singlemicrometer-size fibers from several recombinant silk-like proteins from different spider species, is a powerful tool to generateprotein libraries with corresponding fibers for structure-function relationship investigations in protein-based biomaterials. Thisprotocol may be completed in 40 d.
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