Cloning and expression analysis of alcohol dehydrogenase (Adh) hybrid promoter isolated from Zea mays

Abstract

Hybrid promoters are created by shuffling of DNA fragments while keeping intact regulatory regions crucial of promoter activity. Two fragments of alcohol dehydrogenase (Adh) promoter from Zea mays were selected to generate hybrid promoter. Sequence analysis of both alcohol dehydrogenase promoter fragments through bioinformatics tools identified several crucial cis regulatory elements and transcription factors binding sites. Both fragments were separately cloned in the TA vector (pTZ57R/T) and fused to get the complete hybrid promoter (Adh-H). Alcohol dehydrogenase hybrid promoter was further cloned in expression vector pGR1 through adaptor ligation. Transient β-glucuronidase (GUS) assay revealed that hybrid promoter exhibited high expression under anaerobic conditions in wheat tissues. From the study it is concluded that hybrid promoter (Adh-H) may be used to derive gene expression in monocots during anaerobic conditions. The present work also provides an important insight in the designing of hybrid monocot promoters to improve multiple traits in crops without facing intellectual property rights (IPRs) issues.

Key words: Hybrid promoter, histochemical β-glucuronidase (GUS) assay staining, cis regulatory elements, alcohol dehydrogenase, Zea mays.