Construction and Homologous Expression of a Maize Adh1 Based NcoI Cassette Vector

Abstract

The alcohol dehydrogenase I (Adh1) gene of maize (Zea mays L.) was employed as a source of transcriptional, posttranscriptional, and translational regulatory sequences in the construction of an expression vector. By transforming the translation-initiating ATG and an ATG three triplets upstream from the translational termination triplet into NcoI sites (5'-CCATGG-3'), the maize Adh1 gene was converted into a cassette vector allowing one-step placement of any structural gene under Adh1 regulatory control. We inserted the structural gene for chloramphenicol acetyl transferase (CAT) into this cassette vector and found that this construct expressed the cat gene when transfected into maize protoplasts. Significant expression was observed with a construct that contained only 146 base pairs of Adh1 sequence upstream of the transcription-initiation site. Derivatives with a further 266 or 955 base pairs of contiguous Adh1 upstream sequences increased CAT expression approximately 5-fold or 8-fold, respectively.