Development of a genetic system for a model manganese-oxidizing proteobacterium, Leptothrix discophora SS1
| Autor: | William C. Ghiorse, Leonard W. Lion, Iman A. El Gheriany, Anthony G. Hay, Michael L. Shuler, Daniela Bocioaga |
|---|---|
| Rok vydání: | 2014 |
| Předmět: |
Recombination
Genetic Genetics Manganese biology Discophora Replication Origin biology.organism_classification Origin of replication Microbiology Anti-Bacterial Agents Transformation (genetics) Plasmid Bacterial Proteins Genetic Techniques Conjugation Genetic Leptothrix Leptothrix discophora Homologous recombination Oxidation-Reduction Gene |
| Zdroj: | Microbiology. 160:2396-2405 |
| ISSN: | 1465-2080 1350-0872 |
| Popis: | Understanding the molecular underpinnings of manganese oxidation in Leptothrix discophora SS1 has been hampered by the lack of a genetic system. In this report, we describe the development of a genetic system for L. discophora SS1. The antibiotic sensitivity was characterized, and a procedure for transformation with exogenous DNA via conjugation was developed and optimized, resulting in a maximum transfer frequency of 5.2×10−1 and a typical transfer frequency of the order of 1×10−3 transconjugants per donor. Genetic manipulation of L. discophora SS1 was demonstrated by disrupting pyrF via chromosomal integration with a plasmid containing a R6Kγ origin of replication through homologous recombination. This resulted in resistance to 5-fluoroorotidine, which was abolished by complementation with an ectopically expressed copy of pyrF cloned into pBBR1MCS. This system is expected to be amenable to a systematic genetic analysis of L. discophora SS1, including those genes responsible for manganese oxidation. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|