L‐Ser is often used to synthesize some significant l ‐noncanonical α‐amino acids(l ‐ncAAs), which are the prevalent intermediates and precursors for functional synthetic compounds. In this study, threonine aldolase from Escherichia coli k‐12 MG1655 has been used to synthesize l ‐Ser. In contrast to the maximum catalytic capacity (20 g/L) for l ‐threonine aldolase(LTA), d ‐Ser was synthesized with high yield (240 g/L) from cheap Gly and paraformaldehyde using d ‐threonine aldolase (DTA) from Arthrobacter sp ATCC. In order to fully utilize d ‐Ser and expand the resource of l ‐Ser, a dynamic kinetic resolution system was constructed to convert d /dl ‐Ser to l ‐Ser through combining alanine racemase (Alr) from Bacillus subtilis with l ‐tryptophan synthase (TrpS) from Escherichia coli k‐12 MG1655, and l ‐ncAAs including l ‐Trp and l ‐Cys derivatives were synthesized with excellent enantioselectivity and in high yields. The results indicated l ‐ncAAs could be efficiently synthesized from d ‐Ser using this original and green dynamic kinetic resolution system, and the reliable l ‐Ser resource has been established from simple and achiral substrates.