Full Length Research Paper

Genetic sequences derived from suppression subtractive hybridization analysis provides insight into their possible roles in Xanthomonas albilineans

Rehana Shaik¹, Balakrishna Pillay¹ and Dorsamy Pillay^1,2*

¹Discipline of Microbiology, Faculty of Science and Agriculture, University of KwaZulu-Natal (Westville Campus), Private Bag X54001, Durban 4000, Republic of South Africa.
²Department of Biotechnology, Faculty of Engineering, Science and the Built Environment, Durban University of Technology, P. O. Box 1334, Durban 4000, Republic of South Africa.

*Corresponding author. E-mail: pillayb1@ukzn.ac.za. Tel: +27 312607404. Fax: +27 312607803.

Accepted 4 April, 2008

Leaf scald disease (LSD) is caused by the Gram-negative bacterium, Xanthomonas albilineans. Genomic DNA from X. albilineans and Xanthomonas hyacinthi were analyzed by suppression subtractive hybridization (SSH) using X. albilineans as the tester from which unique sequences were sought and X. hyacinthi as the driver. Following the SSH procedure, amplification products within the size range of 100 - 600 bp were generated, purified, directly cloned with the Promega pGEM-T vector cloning kit, and transformed into ultracompetent Escherichia coli X L2-blue MRF’ cells (Stratagene, La Jolla, CA). Clones selected were sequenced (using a Perkin Elmer ABI PRISM Dye terminator cycle sequencing kit and ABI Model 377 DNA sequencer) in one direction with SP6 and T7 primers (Promega). Clone Xa 6 revealed very close homology with a probable bacterioferritin from Pseudomonas aeruginosa. Clone X. albilineans 12 showed 92% homology to the acetate repressor proteins and clone X. albilineans 18 displayed 85% homology to the plasmid pTOM9 from Alcaligenes xylosoxidans. Sequencing data also revealed homology to various hypothetical proteins.

Key words: suppression subtractive hybridization, Xanthomonas albilineans, Xanthomonas hyacinthi, sequencing.