The Mae I assay for scoring atrazine resistance is codon-usage dependent in legumes

Abstract

Nine different cowpea varieties were grown and total deoxyribonucleic acid (DNA) isolated from the young leaves. This DNA was used to amplify a 291 base pair fragment that contained codon 264 of the psb A gene. The primers used were designed based on the soybean psb A gene sequence. The 291 base pair fragment was digested with the restriction enzyme Mae I to assay for sensitivity/resistance to atrazine using a published method. When Mae I did not cut the 291 base fragment, the whole gene was amplified by the polymerase chain reaction, cloned into pBSK+ll (Stratagene, CA, USA) and sequenced around codon 264. Codons 263 and 264 were found to be GCA and AGT respectively. UWGCG and BLASTsearchesofthepsbAgenesequences in the databases confirmed that Mae I has a recognition site that overlaps with codons 263 and 264 in some legumes but not in others. The Mae I assay used by Cheung and co-workers is therefore not universal but depends on codon usage in legumes.