Methionine is one of the essential amino acids that are synthesized in low amounts in seeds of many agronomically important crop plants such as soybean. Cystathionine-γ-synthase (CS; EC 4.2.99.9) is the branch point enzyme leading to methionine synthesis. We isolated a 1023 nucleotide cDNA-encoding soybean CS from a leaf cDNA library using a 672-nucleotide Arabidopsis cDNA probe. The complete cDNA contains a single open reading frame of 1608 bp that encodes a 536 amino acid protein with a predicted molecular mass of 58 090 Da. The amino terminal portion of the deduced amino acid sequence is rich in threonine and serine, suggesting the presence of a chloroplast transit peptide (155 amino acids). The soybean CS amino acid sequence shares sequence identity with several CS proteins, Arabidopsis thaliana, Z. mays, M. crystallium, and Escherichia coli. The coding region minus the transit peptide was cloned in-frame into pUC18. This construct was used to transform and complement an E. coli methionine auxotroph, AB301. Northern analysis revealed that a 1.9 kb soybean CS mRNA was expressed at the highest level in 8-day light-grown cotyledons and the lowest level in 8-day dark-grown leaves. Southern analysis suggests that the multiple banding patterns may be indicative of several restriction sites within the genomic sequence or CS may be part of a small gene family.