Premethylation of foreign DNA improves integrative transformation efficiency in Synechocystis sp. strain PCC 6803

Bo Wang, Jianping Yu, Weiwen Zhang, Deirdre Meldrum

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

Restriction digestion of foreign DNA is one of the key biological barriers against genetic transformation in microorganisms. To establish a high-efficiency transformation protocol in the model cyanobacterium, Synechocystis sp. strain PCC 6803 (Synechocystis 6803), we investigated the effects of premethylation of foreign DNA on the integrative transformation of this strain. In this study, two type II methyltransferase-encoding genes, i.e., sll0729 (geneM) and slr0214 (gene C), were cloned from the chromosome of Synechocystis 6803 and expressed in Escherichia coli harboring an integration plasmid. After premethylation treatment in E. coli, the integration plasmid was extracted and used for transformation of Synechocystis 6803. The results showed that although expression of methyltransferaseMhad little impact on the transformation of Synechocystis 6803, expression of methyltransferase C resulted in 11- to 161-fold-higher efficiency in the subsequent integrative transformation of Synechocystis 6803. Effective expression of methyltransferase C, which could be achieved by optimizing the 5' untranslated region, was critical to efficient premethylation of the donor DNA and thus high transformation efficiency in Synechocystis 6803. Since premethylating foreign DNA prior to transforming Synechocystis avoids changing the host genetic background, the study thus provides an improved method for high-efficiency integrative transformation of Synechocystis 6803.

Original languageEnglish (US)
Pages (from-to)8500-8506
Number of pages7
JournalApplied and Environmental Microbiology
Volume81
Issue number24
DOIs
StatePublished - 2015

Fingerprint

Synechocystis
DNA
methyltransferases
plasmid
Methyltransferases
plasmids
Phosphatidyl-N-Methylethanolamine N-Methyltransferase
Plasmids
Escherichia coli
Genetic Transformation
gene
5' untranslated regions
genetic transformation
5' Untranslated Regions
Cyanobacteria
genetic background
digestion
cyanobacterium
chromosome
Genes

ASJC Scopus subject areas

  • Applied Microbiology and Biotechnology
  • Food Science
  • Biotechnology
  • Ecology

Cite this

Premethylation of foreign DNA improves integrative transformation efficiency in Synechocystis sp. strain PCC 6803. / Wang, Bo; Yu, Jianping; Zhang, Weiwen; Meldrum, Deirdre.

In: Applied and Environmental Microbiology, Vol. 81, No. 24, 2015, p. 8500-8506.

Research output: Contribution to journalArticle

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