Characterization of the spontaneous elimination of streptomycin sensitivity (Sm^s) on high-copy-number plasmids: Sm^s-enforcement cloning vectors with a synthetic rpsL gene
The strA^s or rpsL^+ gene, encoding a ribosomal protein, S12, expresses its streptomycin-sensitivity (Sm^s) phenotype dominantly over strA^R or rpsL^- gene. Therefore, strA^R cells that harbor plasmids with strA^s alleles are phenotypically Sm^s. It was found that the Sm^s phenotype is unstable, and...
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E-Artikel |
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| Sprache: |
Englisch |
| Erschienen: |
1992 |
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| Reproduktion: |
Elsevier Journal Backfiles on ScienceDirect 1907 - 2002 |
|---|---|
| Übergeordnetes Werk: |
in: Gene - Amsterdam : Elsevier, 121(1992), 1, Seite 25-33 |
| Übergeordnetes Werk: |
volume:121 ; year:1992 ; number:1 ; pages:25-33 |
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NLEJ187308667 |
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| 245 | 1 | 0 | |a Characterization of the spontaneous elimination of streptomycin sensitivity (Sm^s) on high-copy-number plasmids: Sm^s-enforcement cloning vectors with a synthetic rpsL gene |
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| 520 | |a The strA^s or rpsL^+ gene, encoding a ribosomal protein, S12, expresses its streptomycin-sensitivity (Sm^s) phenotype dominantly over strA^R or rpsL^- gene. Therefore, strA^R cells that harbor plasmids with strA^s alleles are phenotypically Sm^s. It was found that the Sm^s phenotype is unstable, and such cells eventually switch to the Sm-resistance (Sm^R) phenotype, especially when the strA^s gene was cloned on high-copy-number (HCN) plasmids. It seemed that the strA gene cloned on HCN plasmids was toxic to Escherichia coli host cells and, during prolonged cultivation, plasmids with an inactivated strA^5 gene, mostly carrying insertion sequence elements, such as IS1, IS5 and γδ, were selected. The instability of the strA gene was particularly enhanced when the Val^5^1 residue in the middle of S12 protein was replaced by Leu, suggesting enhanced toxicity of the altered S12. Since the strA^S gene was stably maintained throughout approx. 100 cell doublings when its expression was abolished, most probably it is the gene product rather than the nucleotide sequence itself that is responsible for the instability of strA gene on HCN plasmids. To improve the stability of the Sm^s phenotype, the previously reported ampicillin-resistance-conferring and Sm^s-enforcing plasmid vector, pHSG670, was reconstructed. The resulting vector, pHSG683, confers chloramphenicol resistance, enforces Sm^s on strA^R and supE^- host bacteria, and has multiple cloning sites within the coding region of synthetic rpsL gene. When pHSG683 DNA was prepared from strA^R and sup^+ cells grown in tryptophan-rich medium with Cm and Sm, less than 10^-^6 plasmids failed to enforce Sm^s on strA^R and supE^- cells in tryptophan-less medium with Cm. This vector is extremely powerful and useful for efficient detection of the rare incidence of DNA recombination or genetic alterations in vitro and in vivo. | ||
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(DE-627)NLEJ187308667 (DE-599)GBVNLZ187308667 DE-627 ger DE-627 rakwb eng Characterization of the spontaneous elimination of streptomycin sensitivity (Sm^s) on high-copy-number plasmids: Sm^s-enforcement cloning vectors with a synthetic rpsL gene 1992 nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier The strA^s or rpsL^+ gene, encoding a ribosomal protein, S12, expresses its streptomycin-sensitivity (Sm^s) phenotype dominantly over strA^R or rpsL^- gene. Therefore, strA^R cells that harbor plasmids with strA^s alleles are phenotypically Sm^s. It was found that the Sm^s phenotype is unstable, and such cells eventually switch to the Sm-resistance (Sm^R) phenotype, especially when the strA^s gene was cloned on high-copy-number (HCN) plasmids. It seemed that the strA gene cloned on HCN plasmids was toxic to Escherichia coli host cells and, during prolonged cultivation, plasmids with an inactivated strA^5 gene, mostly carrying insertion sequence elements, such as IS1, IS5 and γδ, were selected. The instability of the strA gene was particularly enhanced when the Val^5^1 residue in the middle of S12 protein was replaced by Leu, suggesting enhanced toxicity of the altered S12. Since the strA^S gene was stably maintained throughout approx. 100 cell doublings when its expression was abolished, most probably it is the gene product rather than the nucleotide sequence itself that is responsible for the instability of strA gene on HCN plasmids. To improve the stability of the Sm^s phenotype, the previously reported ampicillin-resistance-conferring and Sm^s-enforcing plasmid vector, pHSG670, was reconstructed. The resulting vector, pHSG683, confers chloramphenicol resistance, enforces Sm^s on strA^R and supE^- host bacteria, and has multiple cloning sites within the coding region of synthetic rpsL gene. When pHSG683 DNA was prepared from strA^R and sup^+ cells grown in tryptophan-rich medium with Cm and Sm, less than 10^-^6 plasmids failed to enforce Sm^s on strA^R and supE^- cells in tryptophan-less medium with Cm. This vector is extremely powerful and useful for efficient detection of the rare incidence of DNA recombination or genetic alterations in vitro and in vivo. Elsevier Journal Backfiles on ScienceDirect 1907 - 2002 Toba-Minowa, M. oth Hashimoto-Gotoh, T. oth in Gene Amsterdam : Elsevier 121(1992), 1, Seite 25-33 (DE-627)NLEJ177212578 (DE-600)1491012-3 0378-1119 nnns volume:121 year:1992 number:1 pages:25-33 http://linkinghub.elsevier.com/retrieve/pii/0378-1119(92)90158-L GBV_USEFLAG_H ZDB-1-SDJ GBV_NL_ARTICLE AR 121 1992 1 25-33 |
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(DE-627)NLEJ187308667 (DE-599)GBVNLZ187308667 DE-627 ger DE-627 rakwb eng Characterization of the spontaneous elimination of streptomycin sensitivity (Sm^s) on high-copy-number plasmids: Sm^s-enforcement cloning vectors with a synthetic rpsL gene 1992 nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier The strA^s or rpsL^+ gene, encoding a ribosomal protein, S12, expresses its streptomycin-sensitivity (Sm^s) phenotype dominantly over strA^R or rpsL^- gene. Therefore, strA^R cells that harbor plasmids with strA^s alleles are phenotypically Sm^s. It was found that the Sm^s phenotype is unstable, and such cells eventually switch to the Sm-resistance (Sm^R) phenotype, especially when the strA^s gene was cloned on high-copy-number (HCN) plasmids. It seemed that the strA gene cloned on HCN plasmids was toxic to Escherichia coli host cells and, during prolonged cultivation, plasmids with an inactivated strA^5 gene, mostly carrying insertion sequence elements, such as IS1, IS5 and γδ, were selected. The instability of the strA gene was particularly enhanced when the Val^5^1 residue in the middle of S12 protein was replaced by Leu, suggesting enhanced toxicity of the altered S12. Since the strA^S gene was stably maintained throughout approx. 100 cell doublings when its expression was abolished, most probably it is the gene product rather than the nucleotide sequence itself that is responsible for the instability of strA gene on HCN plasmids. To improve the stability of the Sm^s phenotype, the previously reported ampicillin-resistance-conferring and Sm^s-enforcing plasmid vector, pHSG670, was reconstructed. The resulting vector, pHSG683, confers chloramphenicol resistance, enforces Sm^s on strA^R and supE^- host bacteria, and has multiple cloning sites within the coding region of synthetic rpsL gene. When pHSG683 DNA was prepared from strA^R and sup^+ cells grown in tryptophan-rich medium with Cm and Sm, less than 10^-^6 plasmids failed to enforce Sm^s on strA^R and supE^- cells in tryptophan-less medium with Cm. This vector is extremely powerful and useful for efficient detection of the rare incidence of DNA recombination or genetic alterations in vitro and in vivo. Elsevier Journal Backfiles on ScienceDirect 1907 - 2002 Toba-Minowa, M. oth Hashimoto-Gotoh, T. oth in Gene Amsterdam : Elsevier 121(1992), 1, Seite 25-33 (DE-627)NLEJ177212578 (DE-600)1491012-3 0378-1119 nnns volume:121 year:1992 number:1 pages:25-33 http://linkinghub.elsevier.com/retrieve/pii/0378-1119(92)90158-L GBV_USEFLAG_H ZDB-1-SDJ GBV_NL_ARTICLE AR 121 1992 1 25-33 |
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(DE-627)NLEJ187308667 (DE-599)GBVNLZ187308667 DE-627 ger DE-627 rakwb eng Characterization of the spontaneous elimination of streptomycin sensitivity (Sm^s) on high-copy-number plasmids: Sm^s-enforcement cloning vectors with a synthetic rpsL gene 1992 nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier The strA^s or rpsL^+ gene, encoding a ribosomal protein, S12, expresses its streptomycin-sensitivity (Sm^s) phenotype dominantly over strA^R or rpsL^- gene. Therefore, strA^R cells that harbor plasmids with strA^s alleles are phenotypically Sm^s. It was found that the Sm^s phenotype is unstable, and such cells eventually switch to the Sm-resistance (Sm^R) phenotype, especially when the strA^s gene was cloned on high-copy-number (HCN) plasmids. It seemed that the strA gene cloned on HCN plasmids was toxic to Escherichia coli host cells and, during prolonged cultivation, plasmids with an inactivated strA^5 gene, mostly carrying insertion sequence elements, such as IS1, IS5 and γδ, were selected. The instability of the strA gene was particularly enhanced when the Val^5^1 residue in the middle of S12 protein was replaced by Leu, suggesting enhanced toxicity of the altered S12. Since the strA^S gene was stably maintained throughout approx. 100 cell doublings when its expression was abolished, most probably it is the gene product rather than the nucleotide sequence itself that is responsible for the instability of strA gene on HCN plasmids. To improve the stability of the Sm^s phenotype, the previously reported ampicillin-resistance-conferring and Sm^s-enforcing plasmid vector, pHSG670, was reconstructed. The resulting vector, pHSG683, confers chloramphenicol resistance, enforces Sm^s on strA^R and supE^- host bacteria, and has multiple cloning sites within the coding region of synthetic rpsL gene. When pHSG683 DNA was prepared from strA^R and sup^+ cells grown in tryptophan-rich medium with Cm and Sm, less than 10^-^6 plasmids failed to enforce Sm^s on strA^R and supE^- cells in tryptophan-less medium with Cm. This vector is extremely powerful and useful for efficient detection of the rare incidence of DNA recombination or genetic alterations in vitro and in vivo. Elsevier Journal Backfiles on ScienceDirect 1907 - 2002 Toba-Minowa, M. oth Hashimoto-Gotoh, T. oth in Gene Amsterdam : Elsevier 121(1992), 1, Seite 25-33 (DE-627)NLEJ177212578 (DE-600)1491012-3 0378-1119 nnns volume:121 year:1992 number:1 pages:25-33 http://linkinghub.elsevier.com/retrieve/pii/0378-1119(92)90158-L GBV_USEFLAG_H ZDB-1-SDJ GBV_NL_ARTICLE AR 121 1992 1 25-33 |
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(DE-627)NLEJ187308667 (DE-599)GBVNLZ187308667 DE-627 ger DE-627 rakwb eng Characterization of the spontaneous elimination of streptomycin sensitivity (Sm^s) on high-copy-number plasmids: Sm^s-enforcement cloning vectors with a synthetic rpsL gene 1992 nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier The strA^s or rpsL^+ gene, encoding a ribosomal protein, S12, expresses its streptomycin-sensitivity (Sm^s) phenotype dominantly over strA^R or rpsL^- gene. Therefore, strA^R cells that harbor plasmids with strA^s alleles are phenotypically Sm^s. It was found that the Sm^s phenotype is unstable, and such cells eventually switch to the Sm-resistance (Sm^R) phenotype, especially when the strA^s gene was cloned on high-copy-number (HCN) plasmids. It seemed that the strA gene cloned on HCN plasmids was toxic to Escherichia coli host cells and, during prolonged cultivation, plasmids with an inactivated strA^5 gene, mostly carrying insertion sequence elements, such as IS1, IS5 and γδ, were selected. The instability of the strA gene was particularly enhanced when the Val^5^1 residue in the middle of S12 protein was replaced by Leu, suggesting enhanced toxicity of the altered S12. Since the strA^S gene was stably maintained throughout approx. 100 cell doublings when its expression was abolished, most probably it is the gene product rather than the nucleotide sequence itself that is responsible for the instability of strA gene on HCN plasmids. To improve the stability of the Sm^s phenotype, the previously reported ampicillin-resistance-conferring and Sm^s-enforcing plasmid vector, pHSG670, was reconstructed. The resulting vector, pHSG683, confers chloramphenicol resistance, enforces Sm^s on strA^R and supE^- host bacteria, and has multiple cloning sites within the coding region of synthetic rpsL gene. When pHSG683 DNA was prepared from strA^R and sup^+ cells grown in tryptophan-rich medium with Cm and Sm, less than 10^-^6 plasmids failed to enforce Sm^s on strA^R and supE^- cells in tryptophan-less medium with Cm. This vector is extremely powerful and useful for efficient detection of the rare incidence of DNA recombination or genetic alterations in vitro and in vivo. Elsevier Journal Backfiles on ScienceDirect 1907 - 2002 Toba-Minowa, M. oth Hashimoto-Gotoh, T. oth in Gene Amsterdam : Elsevier 121(1992), 1, Seite 25-33 (DE-627)NLEJ177212578 (DE-600)1491012-3 0378-1119 nnns volume:121 year:1992 number:1 pages:25-33 http://linkinghub.elsevier.com/retrieve/pii/0378-1119(92)90158-L GBV_USEFLAG_H ZDB-1-SDJ GBV_NL_ARTICLE AR 121 1992 1 25-33 |
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(DE-627)NLEJ187308667 (DE-599)GBVNLZ187308667 DE-627 ger DE-627 rakwb eng Characterization of the spontaneous elimination of streptomycin sensitivity (Sm^s) on high-copy-number plasmids: Sm^s-enforcement cloning vectors with a synthetic rpsL gene 1992 nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier The strA^s or rpsL^+ gene, encoding a ribosomal protein, S12, expresses its streptomycin-sensitivity (Sm^s) phenotype dominantly over strA^R or rpsL^- gene. Therefore, strA^R cells that harbor plasmids with strA^s alleles are phenotypically Sm^s. It was found that the Sm^s phenotype is unstable, and such cells eventually switch to the Sm-resistance (Sm^R) phenotype, especially when the strA^s gene was cloned on high-copy-number (HCN) plasmids. It seemed that the strA gene cloned on HCN plasmids was toxic to Escherichia coli host cells and, during prolonged cultivation, plasmids with an inactivated strA^5 gene, mostly carrying insertion sequence elements, such as IS1, IS5 and γδ, were selected. The instability of the strA gene was particularly enhanced when the Val^5^1 residue in the middle of S12 protein was replaced by Leu, suggesting enhanced toxicity of the altered S12. Since the strA^S gene was stably maintained throughout approx. 100 cell doublings when its expression was abolished, most probably it is the gene product rather than the nucleotide sequence itself that is responsible for the instability of strA gene on HCN plasmids. To improve the stability of the Sm^s phenotype, the previously reported ampicillin-resistance-conferring and Sm^s-enforcing plasmid vector, pHSG670, was reconstructed. The resulting vector, pHSG683, confers chloramphenicol resistance, enforces Sm^s on strA^R and supE^- host bacteria, and has multiple cloning sites within the coding region of synthetic rpsL gene. When pHSG683 DNA was prepared from strA^R and sup^+ cells grown in tryptophan-rich medium with Cm and Sm, less than 10^-^6 plasmids failed to enforce Sm^s on strA^R and supE^- cells in tryptophan-less medium with Cm. This vector is extremely powerful and useful for efficient detection of the rare incidence of DNA recombination or genetic alterations in vitro and in vivo. Elsevier Journal Backfiles on ScienceDirect 1907 - 2002 Toba-Minowa, M. oth Hashimoto-Gotoh, T. oth in Gene Amsterdam : Elsevier 121(1992), 1, Seite 25-33 (DE-627)NLEJ177212578 (DE-600)1491012-3 0378-1119 nnns volume:121 year:1992 number:1 pages:25-33 http://linkinghub.elsevier.com/retrieve/pii/0378-1119(92)90158-L GBV_USEFLAG_H ZDB-1-SDJ GBV_NL_ARTICLE AR 121 1992 1 25-33 |
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characterization of the spontaneous elimination of streptomycin sensitivity (sm^s) on high-copy-number plasmids: sm^s-enforcement cloning vectors with a synthetic rpsl gene |
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Characterization of the spontaneous elimination of streptomycin sensitivity (Sm^s) on high-copy-number plasmids: Sm^s-enforcement cloning vectors with a synthetic rpsL gene |
| abstract |
The strA^s or rpsL^+ gene, encoding a ribosomal protein, S12, expresses its streptomycin-sensitivity (Sm^s) phenotype dominantly over strA^R or rpsL^- gene. Therefore, strA^R cells that harbor plasmids with strA^s alleles are phenotypically Sm^s. It was found that the Sm^s phenotype is unstable, and such cells eventually switch to the Sm-resistance (Sm^R) phenotype, especially when the strA^s gene was cloned on high-copy-number (HCN) plasmids. It seemed that the strA gene cloned on HCN plasmids was toxic to Escherichia coli host cells and, during prolonged cultivation, plasmids with an inactivated strA^5 gene, mostly carrying insertion sequence elements, such as IS1, IS5 and γδ, were selected. The instability of the strA gene was particularly enhanced when the Val^5^1 residue in the middle of S12 protein was replaced by Leu, suggesting enhanced toxicity of the altered S12. Since the strA^S gene was stably maintained throughout approx. 100 cell doublings when its expression was abolished, most probably it is the gene product rather than the nucleotide sequence itself that is responsible for the instability of strA gene on HCN plasmids. To improve the stability of the Sm^s phenotype, the previously reported ampicillin-resistance-conferring and Sm^s-enforcing plasmid vector, pHSG670, was reconstructed. The resulting vector, pHSG683, confers chloramphenicol resistance, enforces Sm^s on strA^R and supE^- host bacteria, and has multiple cloning sites within the coding region of synthetic rpsL gene. When pHSG683 DNA was prepared from strA^R and sup^+ cells grown in tryptophan-rich medium with Cm and Sm, less than 10^-^6 plasmids failed to enforce Sm^s on strA^R and supE^- cells in tryptophan-less medium with Cm. This vector is extremely powerful and useful for efficient detection of the rare incidence of DNA recombination or genetic alterations in vitro and in vivo. |
| abstractGer |
The strA^s or rpsL^+ gene, encoding a ribosomal protein, S12, expresses its streptomycin-sensitivity (Sm^s) phenotype dominantly over strA^R or rpsL^- gene. Therefore, strA^R cells that harbor plasmids with strA^s alleles are phenotypically Sm^s. It was found that the Sm^s phenotype is unstable, and such cells eventually switch to the Sm-resistance (Sm^R) phenotype, especially when the strA^s gene was cloned on high-copy-number (HCN) plasmids. It seemed that the strA gene cloned on HCN plasmids was toxic to Escherichia coli host cells and, during prolonged cultivation, plasmids with an inactivated strA^5 gene, mostly carrying insertion sequence elements, such as IS1, IS5 and γδ, were selected. The instability of the strA gene was particularly enhanced when the Val^5^1 residue in the middle of S12 protein was replaced by Leu, suggesting enhanced toxicity of the altered S12. Since the strA^S gene was stably maintained throughout approx. 100 cell doublings when its expression was abolished, most probably it is the gene product rather than the nucleotide sequence itself that is responsible for the instability of strA gene on HCN plasmids. To improve the stability of the Sm^s phenotype, the previously reported ampicillin-resistance-conferring and Sm^s-enforcing plasmid vector, pHSG670, was reconstructed. The resulting vector, pHSG683, confers chloramphenicol resistance, enforces Sm^s on strA^R and supE^- host bacteria, and has multiple cloning sites within the coding region of synthetic rpsL gene. When pHSG683 DNA was prepared from strA^R and sup^+ cells grown in tryptophan-rich medium with Cm and Sm, less than 10^-^6 plasmids failed to enforce Sm^s on strA^R and supE^- cells in tryptophan-less medium with Cm. This vector is extremely powerful and useful for efficient detection of the rare incidence of DNA recombination or genetic alterations in vitro and in vivo. |
| abstract_unstemmed |
The strA^s or rpsL^+ gene, encoding a ribosomal protein, S12, expresses its streptomycin-sensitivity (Sm^s) phenotype dominantly over strA^R or rpsL^- gene. Therefore, strA^R cells that harbor plasmids with strA^s alleles are phenotypically Sm^s. It was found that the Sm^s phenotype is unstable, and such cells eventually switch to the Sm-resistance (Sm^R) phenotype, especially when the strA^s gene was cloned on high-copy-number (HCN) plasmids. It seemed that the strA gene cloned on HCN plasmids was toxic to Escherichia coli host cells and, during prolonged cultivation, plasmids with an inactivated strA^5 gene, mostly carrying insertion sequence elements, such as IS1, IS5 and γδ, were selected. The instability of the strA gene was particularly enhanced when the Val^5^1 residue in the middle of S12 protein was replaced by Leu, suggesting enhanced toxicity of the altered S12. Since the strA^S gene was stably maintained throughout approx. 100 cell doublings when its expression was abolished, most probably it is the gene product rather than the nucleotide sequence itself that is responsible for the instability of strA gene on HCN plasmids. To improve the stability of the Sm^s phenotype, the previously reported ampicillin-resistance-conferring and Sm^s-enforcing plasmid vector, pHSG670, was reconstructed. The resulting vector, pHSG683, confers chloramphenicol resistance, enforces Sm^s on strA^R and supE^- host bacteria, and has multiple cloning sites within the coding region of synthetic rpsL gene. When pHSG683 DNA was prepared from strA^R and sup^+ cells grown in tryptophan-rich medium with Cm and Sm, less than 10^-^6 plasmids failed to enforce Sm^s on strA^R and supE^- cells in tryptophan-less medium with Cm. This vector is extremely powerful and useful for efficient detection of the rare incidence of DNA recombination or genetic alterations in vitro and in vivo. |
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| title_short |
Characterization of the spontaneous elimination of streptomycin sensitivity (Sm^s) on high-copy-number plasmids: Sm^s-enforcement cloning vectors with a synthetic rpsL gene |
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Toba-Minowa, M. Hashimoto-Gotoh, T. |
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