Nebulin is an approximately 700 kDa filamentous protein in vertebrate skeletal muscle. It binds to the Z line and also binds side-by-side to the entire thin (actin) filament in a sarcomere. Nebulin is currently thought to be a molecular ruler regulating the length of the thin filament to 1 mum. The complete sequence of human skeletal muscle nebulin was determined by . Because of its large size, only fragmental sequence information has been available for nebulins other than human skeletal muscle. This paper describes for the first time the sequence of about one third (C terminal region) of chicken skeletal muscle nebulin. It was found that the fundamental structure of human nebulin, consisting of 35 amino acid repeats (modules) plus C terminal serine-rich and SH3 domains linked to the Z line are well conserved with chicken nebulin. Sequence identity ranged from 74 to 91%. There were super-repeats (seven modules), a first linker repeat, simple repeat and a second linker repeat in addition to the Z line binding region as in human nebulin. However, there were 2 fewer modules in the first linker repeat and 6 fewer in the simple repeat in chicken nebulin as compared to human nebulin. Two isoforms of chicken nebulin were sequenced indicating insertion of approximately 6 or 11 modules to a structure similar to that of human nebulin. Recombinant first linker repeats M51 approximately 56 were shown to bind to actin using the ELISA technique as well as human nebulin recombinants.