Isolation and cloning of cytokinin oxidase 5 gene from Arabidopsis

Abstract

Phytohormones play intricate physiological and biochemical roles to control growth and
development of plants. Amongst several phytohormones cytokinins (CKs) play an important role
as they govern yield in crops. While controlling cardinal growth of plants, CKs govern a complex
trait i.e. shoot apical meristem formation and there by yield in crops. Mutation studies on cytokinin
oxidase (ckx) genes had revealed that decreased activity of cytokinin oxidase results in increased
bioavailability of cytokinin and thereby increasing seed yield. In this study, we isolated ckx5
gene from Arabidopsis and cloned into an E coli vector so as to make it ready for mobilization into
binary vector for future plant transformation experiments. Sequence of steps followed to clone
the gene from Arabidopsis and confirm its presence were ncomphishel by (i) PCR based amplification
with gene specific primers, (ii) restriction digestion followed by (iii) authentication of ckx gene by
Sanger's DNA-sequencing are discussed.