characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2022-02-09T16:22:06Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2022-02-09T16:22:06Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA10166569
characteristics.characteristic: External Id
characteristics.text: Centre for Genomic Regulation
characteristics.characteristic: INSDC center name
characteristics.text: 2022-02-09T16:22:06Z
characteristics.characteristic: INSDC first public
characteristics.text: 2022-02-09T16:22:06Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS7823045
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-10981:C5_P15_R2
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: Protocols: The clonal strains C5, C20 and C26 used in this study are derived from M129 wild type strain (ATCC 29342) and were cultivated in SP-4 medium with antibiotics and inducers when appropriate, at 37ºC and 5% CO2. The strain C5 has a single minitransposon with the KS1 kill-switch inserted into the gene mpn462 gene. The strain C20, derived from strain C5, has a second minitransposon with the KS13g kill-switch inserted in the intergenic region between genes mpn648 and mpn649. And, the strain C26, also derived from strain C5, has a second minitransposon with the TKS2 kill-switch inserted into the mpn062 gene. All strains C5, C20 and C26 were serially passaged every 48h for 15th passages in SP-4 medium supplemented with puromycin (3.3 µg/mL), puromycin and chloramphenicol (20 µg/mL), or puromycin, chloramphenicol and theophylline (0.5 mM), respectively. Genomic DNA was isolated from samples of strain the strains C5 and C20 grown in the permissive condition (in the absence of IPTG) at the 2nd, 3rd and 15th passage. Genomic DNA was also isolated from samples of the strain C26 at passages 2nd and 15th grown in permissive conditions with 0.5 mM theophylline. In the case of the strain C5, genomic DNA was also isolated from a sample of the strain at the 3rd passage grown in the presence of 1 mM IPTG that triggers the kill-switch present in the strain. PX in the name of the samples correspond to the number of passages applied IPTG kill-switch induction in samples C5_P3IPTG Genomic DNA samples were prepared from late-exponential phase cultures of M. pneumoniae strains C5, C20 and C26 grown in SP-4 medium (and relevant antibiotics and inducers). Cells grown in 75 cm2 flasks were washed three times with PBS and followed by genomic extraction using the GenElute™ Bacterial Genomic DNA Kit (Cat. No. NA2110, Sigma). The London Biofoundry (Imperial College London) performed sequencing: 500 ng of the Extracted DNA genomes were prepared for sequencing with the Illumina Nextera DNA Flex kit (20018704) according to manufacturer recommendations via an automated implementation on an Opentrons OT-2 liquid handler.
characteristics.characteristic: description
characteristics.text: KS1 kill-switch
characteristics.characteristic: genotype
characteristics.text: Mycoplasma pneumoniae M129
characteristics.characteristic: organism
characteristics.text: Mycoplasma pneumoniae M129
characteristics.characteristic: organism
characteristics.text: whole organism
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/UBERON_0000468
characteristics.characteristic: organism part
characteristics.text: P15
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0002948
characteristics.characteristic: passage
characteristics.text: 2
characteristics.characteristic: replicate
characteristics.text: Mycoplasma pneumoniae M129
characteristics.characteristic: scientific_name
characteristics.text: C5_P15_R2
characteristics.characteristic: title