Molecular cloning and sequence comparison of a cDN

Authors: Anshu Bansal Niranjan Das

Publish Date: 2012/11/15

Volume: 22, Issue: 4, Pages: 441-452

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Abstract

Starch phosphorylation is an important biochemical aspect of plant starch metabolism as it influences the overall structure of the starch granule and a prerequisite for its degradation There is a growing interest on the isolation and characterization of αglucan/glucanlike water dikinases GWDs from plants particularly agriculturally important crops because GWD is known to catalyze starch phosphorylation both in leaves and different plant storage organs In the present study a 4789bp fulllength cDNA encoding a GWD isoform was isolated from a commercially important Indian potato cultivar Kufri Chipsona1 by RTPCR approach using tuber RNA The predicted protein consisted of 1463 amino acids having Nterminal 77amino acid transit peptide and 1386amino acid mature protein shorter by one amino acid as compared to the other mature GWDs from potato and tomato The mature GWD showed 98  sequence identity with the GWD isolated earlier from the potato cv Desiree Variations were found at 25 locations representing mostly nonconservative substitutions The GWD represents a distinct isoform from potato as revealed by sequence and phylogenetic analyses Amino acid composition segmentwise hydrophobic characters predicted secondary structures were also analyzed and documented in this report Broadly the level of GWD expression as analyzed by semiquantitative RTPCR approach was found to be nearly uniform both in the mature tubers and leaves from most of the potato cultivars By immunodetection technique a band corresponding to ~155 kDa protein was detected only in the tuber protein extracts The tuber starchbound phosphorus content data showed minor variations between the potato cultivars

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