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A PCR technique based on the Hipl interspersed repetitive sequence distinguishes cyanobacterial species and strains

¹IENS, Division of Biological Sciences, Lancaster University, Bailrigg, Lancaster LA1 4YQ, UK
²Culture Collection of Algae and Protozoa (CCAP), IFE, Windermere Laboratory, The Ferry House, Far Sawrey, Ambleside, Cumbria LA22 OLP, UK

¹ Author for correspondence: R. J. Smith. Tel: +44 1524 65201 ext. 93515. Fax: +44 1524 843854. e-mail: r.smith@lancaster.ac.uk

ABSTRACT

SUMMARY: The use of primers based on the Hipl sequence as a typing technique for cyanobacteria has been investigated. The discovery of short repetitive sequence structures in bacterial DNA during the last decade has led t o the development of PCR-based methods for typing, i.e. distinguishing and identifying, bacterial species and strains. An octameric palindromic sequence known as Hipl has been shown t o be present in the chromosomal DNA of many species of cyanobacteria as a highly repetitious interspersed sequence. PCR primers were constructed that extended the Hipl sequence at the 3` end by two bases. Five of the 16 possible extended primers were tested. Each of the five primers produced a different set of products when used t o prime PCR from cyanobacterial genomic DNA. Each primer produced a distinct set of products for each of the 15 cyanobacterial species tested. The ability of Hipl-based PCR to resolve taxonomic differences was assessed by analysis of independent isolates of Anabaena flos-aquae and Nostoc ellipsosporum obtained from the CCAP (Culture Collection of Algae and Protozoa, IFE, Cumbria, UK). A PCR-based RFLP analysis of products amplified from the 235-165 rDNA intergenic region was used to characterize the isolates and to compare with the Hipl typing data. The RFLP and Hipl typing yielded similar results and both techniques were able to distinguish different strains. On the basis of these results it is suggested that the Hipl PCR technique may assist in distinguishing cyanobacterial species and strains.

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