Summary
Synechocystis 6803 contains at least four cryptic plasmids of 2.27 kb (pUS1, pUS2 and pUS3) and 5.20 kb (pUS4). The 1.70 kb HpaI fragments of the related plasmids pUS2 and pUS3 were cloned into the Apr gene of the E. coli plasmid pACYC177, yielding the Kmr hybrid plasmids pUF12 and pUF3 respectively. pUF3 recombines in Synechocystis 6803 with a 2.27 kb plasmid giving the Kmr shuttle vector pUF311. The 1.35 kb HaeII fragment containing the Cm2 gene of the E. coli plasmid pACYC184 was cloned in pUF311 generating the Cmr Kmr shuttle vector pFCLV7. Wild-type cells of Synechocystis 6803 are transformed, albeit poorly, by the plasmids pUF3, pUF12 and pFCLV7. pFCLV7 very efficiently transforms the SUF311 strain of Synechocystis 6803 containing pUF311 as a resident plasmid. This is due to recombination between the homologous parts of pFCLV7 and pUF311. For the same reason the strain SUF311 is also efficiently transformable by E. coli plasmids, as shown for pLF8, provided that they have some homology with the E. coli part of pUF311.
The combined use of Synechocystis 6803 strain SUF311 and of plasmids pFCLV7 and pLF8 generates an efficient host-vector system for gene cloning in this facultatively heterotrophic cyanobacterium.
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References
Astier C, Espardellier F (1976) Mise en evidence d'un systeme de transfert genetique chez une cyanophycee du genre Aphanocapsa. Compt Rend Acad Sci Paris T 282:795–797
Astier C, Elmorjani K, Meyer I, Joset F, Herdman M (1984) Photosynthetic mutants of the cyanobacteria Synechocystis Sp. strains PCC 6714 and PCC 6803: sodium p-hydroxymercuribenzoate as a selective agent. J Bacteriol 158:659–664
Chang ACY, Cohen SN (1978) Construction and characterization of amplifiable multicopy DNA cloning vehicles derived from the p15A cryptic mini plasmid. J Bacteriol 134:1141–1156
Chauvat F, Astier C, Vedel F, Joset-Espardellier F (1983) Transformation in the Cyanobacterium Synechococcus R2: Improvement of efficiency; role of the pUH24 plasmid. Mol Gen Genet 191:39–45
Cohen SN, Chang ACY, Hsu L (1972) Nonchromosomal antibiotic resistance in bacteria: Genetic transformation of Escherichia-coli by R-factor DNA. Proc Natl Acad Sci USA 69:2110–2114
De Vos WM, Venema G, Canosi U, Trautner TA (1981) Plasmid transformation in Bacillus subtilis: fate of plasmid DNA. Mol Gen Genet 181:424–433
Grigorieva G, Shestakov S (1982) Transformation in the cyanobacterium Synechocystis sp 6803. FEMS Microbiol Lett 13:367–370
Kuhlemeier CJ, Borrias WE, Van den Hondel CAMJJ, Van Arkel GA (1981) Vectors for cloning in cyanobacteria: construction and characterization of two recombinant plasmids capable of transformation to Escherichia coli K12 and Anacystis nidulans R2. Mol Gen Genet 184:249–254
Kuhlemeier CJ, Thomas AAM, Van der Ende A, Van Leen RW, Borrias WE, Van den Hondel CAMJJ, Van Arkel GA (1983) A host-vector system for gene cloning in the cyanobacterium Anacystis nidulans R2. Plasmid 10:156–163
Kuhlemeier CJ, Logtenberg T, Stoorvogel W, Van Heugten HAA, Borrias WE, Van Arkel GA (1984a) Cloning of nitrate reductase genes from the cyanobacterium Anacystis nidulans. J Bacteriol 159:36–41
Kuhlemeier CJ, Teeuwsen VJP, Janssen MJT, Van Arkel GA (1984b) Cloning of a third nitrate reductase gene from the cyanobacterium Anacystis nidulans R2 using a shuttle cosmid library. Gene 31:109–116
Lambert GR, Scott JG, Carr NG (1984) Characterization and cloning of extrachromosomal DNA from filamentous cyanobacteria. FEMS Microbiol Lett 21:225–231
Lau RH, Doolittle WF (1979) Covalently closed circular DNAs in closely related unicellular cyanobacteria. J Bacteriol 137:648–652
Maniatis T, Fritsch EF, Sambrook J (1982) molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory, pp 366–369
Parker RC, Watson RM, Vinograd J (1977) Mapping of closed circular DNAs by cleavage with restriction endonucleases and calibration by agarose gel electrophoresis. Proc Natl Acad Sci USA 74:851–855
Potts M (1984) Distribution of plasmids in cyanobacteria of the LPP group. FEMS Microbiol Lett 24:351–354
Rippka R, Deruelles J, Waterbury JB, Herdman M, Stanier RY (1979) Generic assignments, strain histories and properties of pure cultures of cyanobacteria. J Gen Microbiol 111:1–61
Simon RD (1978) Survey of extrachromosomal DNA found in the filamentous cyanobacteria. J Bacteriol 136:414–418
Stüber D, Bujard H (1981) Organization of transcriptional signals in plasmids pBR322 and pACYC184. Proc Natl Acad Sci USA 78:167–171
Tandeau de Marsac N, Borrias WE, Kuhlemeier CJ, Castets AM, Van Arkel GA, Van den Hondel CAMJJ (1982) A new approach for molecular cloning in cyanobacteria: cloning of an Anacystis nidulans met gene using a Tn901-induced mutant. Gene 20:111–119
Van den Hondel CAMJJ, Keegstra W, Borrias WE, Van Arkel GA (1979) Homology of plasmids in strains of unicellular cyanobacteria. Plasmid 2:323–333
Yang RCA, Lis J, Wu R (1979) Elution of DNA from agarose gels after electrophoresis. Methods Enzymol 68:176–182
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Communicated by J.W. Lengeler
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Chauvat, F., De Vries, L., Van der Ende, A. et al. A host-vector system for gene cloning in the cyanobacterium Synechocystis PCC 6803 . Molec Gen Genet 204, 185–191 (1986). https://doi.org/10.1007/BF00330208
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DOI: https://doi.org/10.1007/BF00330208