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Product Name:
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A909 (Group B) Transposon Mutant Library, Plate 12
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Manufacturer:
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BEI Resources
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Additional Information:
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Production in the 96-well format has increased risk of cross-contamination between adjacent wells. Individual clones should be purified (e.g., single colony isolation and purification using good microbiological practices) and sequence-verified prior to use. BEI Resources does not confirm or validate individual mutants provided by the contributor.
The Streptococcus agalactiae, Strain A909 (group B) indexed transposon mutant library consists of 21 plates containing approximately 1900 sequence-defined Himar1 mini-transposon mutant strains, each with a unique disruption to its chromosome. The Himar1 mini-transposon inserts at random TA dinucleotide sites throughout the Group B Streptococcus (GBS) genome, resulting in interruptions in genes and intergenic regions representing a wide range of predicted functional categories, including roles in metabolism, structure and virulence.1,2
Mutagenesis occurred through the introduction and subsequent curing of the temperature-sensitive shuttle vector pCAM48, containing the 1,455 base pair Himar1 mini-transposon, flanked by terminal inverted repeat sequences in which the MmeI restriction enzyme sites required for transposon insertion sequencing are integrated. An erythromycin (Erm) resistance marker, R6kγ origin of replication (Ori R6κγ) for plasmid rescue, and the C9 Himar1 transposase gene, under the control of the S. pyogenes M1 gyrA promoter, required for transposon insertion at random TA dinucleotide sites, are included within the transposon. Outside of the transposon, pCAM48 contains a ColE1 origin of replication; the Gram-positive, temperature-sensitive replicase RepA TS, in which the single base pair deletion present in the progenitor plasmid pCAM45 was repaired; and a kanamycin resistance marker (Kanr).1,2,3 Detailed information about each mutant is shown in Table 1.
Plate orientation and viability were confirmed for NR-59489.
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Material Provided:
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Each inoculated well of the 96-well plate contains approximately 60 µL of culture in Todd-Hewitt broth containing 5 µg/mL erythromycin supplemented with 10% glycerol.
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Packing/Storage:
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NR-59489 was packaged aseptically in a 96-well plate. The product is provided frozen and should be stored at -80°C or colder immediately upon arrival. For long-term storage, the vapor phase of a liquid nitrogen freezer is recommended. Freeze-thaw cycles should be avoided.
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Growth Conditions:
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Media:
Todd-Hewitt broth containing 5 µg/mL erythromycin or equivalent
CHROMagar™ StrepB agar containing 5 µg/mL erythromycin or equivalent
Note: Growth in erythromycin-containing selective media is not required for the propagation of mutants from this library.1
Incubation:
Temperature: 37°C
Atmosphere: Aerobic with 5% CO2
Propagation:
1. Scrape top of frozen well with a pipette tip and streak onto agar plate.
2. Incubate the plates at 37°C for 1 to 2 days.
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Disclaimers:
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You are authorized to use this product for research use only. It is not intended for human use. Use of this product is subject to the terms and conditions of the BEI Resources Material Transfer Agreement (MTA). The MTA is available on our Web site at www.beiresources.org. While BEI Resources uses reasonable efforts to include accurate and up-to-date information on this product sheet, neither ATCC® nor the U.S. Government makes any warranties or representations as to its accuracy. Citations from scientific literature and patents are provided for informational purposes only. Neither ATCC® nor the U.S. Government warrants that such information has been confirmed to be accurate. This product is sent with the condition that you are responsible for its safe storage, handling, use and disposal. ATCC® and the U.S. Government are not liable for any damages or injuries arising from receipt and/ or use of this product. While reasonable effort is made to ensure authenticity and reliability of materials on deposit, the U.S. Government, ATCC®, their suppliers and contributors to BEI Resources are not liable for damages arising from the misidentification or misrepresentation of products. |
References:
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1. Bhavana, B. H., et al. “A Group B Streptococcus Indexed Transposon Mutant Library to Accelerate Genetic Research on an Important Perinatal Pathogen.” Microbiol. Spectr. 11 (2023): e0204623. PubMed: 37933989.
2. Hooven, T. A., et al. “The Essential Genome of Streptococcus agalactiae.” BMC Genomics 17 (2016): 406. PubMed: 27229469.
3. Hooven, T. A., et al. “The Streptococcus agalactiae Stringent Response Enhances Virulence and Persistence in Human Blood.” Infect. Immun. 86 (2017): e00612-17. PubMed: 29109175.
4. Dammann, A. N., et al. “Genome-Wide Fitness Analysis of Group B Streptococcus in Human Amniotic Fluid Reveals a Transcription Factor that Controls Multiple Virulence Traits.” PLoS Patho. 17 (2021): e1009116. PubMed: 33684178.
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Citation:
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Acknowledgment for publications should read “The following reagent was obtained through BEI Resources, NIAID, NIH: Streptococcus agalactiae, Strain A909 (Group B), Transposon Mutant Library, Plate 12, NR-59489.”
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Biosafety Level:
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2
Appropriate safety procedures should always be used with this material. Laboratory safety is discussed in the following publication: U.S. Department of Health and Human Services, Public Health Service, Centers for Disease Control and Prevention, and National Institutes of Health. Biosafety in Microbiological and Biomedical Laboratories (BMBL). Current Edition. Washington, DC: U.S. Government Printing Office.
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