Production and phenotypic analysis of rice transgenics with altered levels of pyruvate decarboxylase and alcohol dehydrogenase proteins

Abstract

Pyruvate decarboxylase (Pdc) and alcohol dehydrogenase (Adh) enzymes are responsible for the operation of ethanolic fermentation pathway that appears to correlate to an extent with anoxia tolerance in plants. This study was undertaken with the objective of (a) analysing the rice pdc gene family and (b) altering the efficacy of the ethanolic fermentation process, through production of transgenic rice plants over- and under-expressing pyruvate decarboxylase (employing Ospdc1 gene from rice) as well as over-expressing alcohol dehydrogenase (employing Ghadh2 gene from cotton) proteins. Correlations noted in this study between the pattern of expression of the Pdc α-subunit and Ospdc2 transcript as well as between the Pdc β-subunit and Ospdc1 transcript suggest the possibility that α-subunit is encoded by Ospdc2 and that β-subunit is encoded by Ospdc1. The fact that levels of Pdc β-subunit were particularly high in pUH-sPdc1 (plasmid construct designed for over-expression of Ospdc1) seedlings while levels of β-subunit levels were negligible or lower in pUH-asPdc1 (plasmid construct designed for under-expression of Ospdc1) seedlings also support these observations. Transgenics raised for over-expression of Pdc and Adh and under-expression of Pdc were confirmed for the transgene presence and effects by PCR, Southern blotting, Northern blotting, Western blotting and isozyme assays. Pdc and Adh over-expressing rice transgenics at early seedling stage under unstressed control growth conditions showed slight, consistent advantage in root vigour as compared to that of wild-type seedlings.